Development of model observers applied to 3D breast tomosynthesis microcalcifications and masses

The development of model observers for mimicking human detection strategies has followed from symmetric signals in simple noise to increasingly complex backgrounds. In this study we implement different model observers for the complex task of detecting a signal in a 3D image stack. The backgrounds come from real breast tomosynthesis acquisitions and the signals were simulated and reconstructed within the volume. Two different tasks relevant to the early detection of breast cancer were considered: detecting an 8 mm mass and detecting a cluster of microcalcifications. The model observers were calculated using a channelized Hotelling observer (CHO) with dense difference-of-Gaussian channels, and a modified (Partial prewhitening [PPW]) observer which was adapted to realistic signals which are not circularly symmetric. The sustained temporal sensitivity function was used to filter the images before applying the spatial templates. For a frame rate of five frames per second, the only CHO that we calculated performed worse than the humans in a 4-AFC experiment. The other observers were variations of PPW and outperformed human observers in every single case. This initial frame rate was a rather low speed and the temporal filtering did not affect the results compared to a data set with no human temporal effects taken into account. We subsequently investigated two higher speeds at 5, 15 and 30 frames per second. We observed that for large masses, the two types of model observers investigated outperformed the human observers and would be suitable with the appropriate addition of internal noise. However, for microcalcifications both only the PPW observer consistently outperformed the humans. The study demonstrated the possibility of using a model observer which takes into account the temporal effects of scrolling through an image stack while being able to effectively detect a range of mass sizes and distributions.

Prox1 interacts with Atoh1 and Gfi1, and regulates cellular differentiation in the inner ear sensory epithelia.

Inner ear hair cells and supporting cells arise from common precursors and, in mammals, do not show phenotypic conversion. Here, we studied the role of the homeodomain transcription factor Prox1 in the inner ear sensory epithelia. Adenoviral-mediated Prox1 transduction into hair cells in explant cultures led to strong repression of Atoh1 and Gfi1, two transcription factors critical for hair cell differentiation and survival. Luciferase assays showed that Prox1 can repress transcriptional activity of Gfi1 independently of Atoh1. Prox1 transduction into cochlear outer hair cells resulted in degeneration of these cells, consistent with the known phenotype of Gfi1-deficient mice. These results together with the widespread expression of endogenous Prox1 within the population of inner ear supporting cells point to the role for Prox1 in antagonizing the hair cell phenotype in these non-sensory cells. Further, in vivo analyses of hair cells from Gfi1-deficient mice suggest that the cyclin-dependent kinase inhibitor p57(Kip2) mediates the differentiation- and survival-promoting functions of Gfi1. These data reveal novel gene interactions and show that these interactions regulate cellular differentiation within the inner ear sensory epithelia. The data point to the tight regulation of phenotypic characteristics of hair cells and supporting cells.

Spatio-temporal differentiation and sociality in spiders.

Species that differ in their social system, and thus in traits such as group size and dispersal timing, may differ in their use of resources along spatial, temporal, or dietary dimensions. The role of sociality in creating differences in habitat use is best explored by studying closely related species or socially polymorphic species that differ in their social system, but share a common environment. Here we investigate whether five sympatric Anelosimus spider species that range from nearly solitary to highly social differ in their use of space and in their phenology as a function of their social system. By studying these species in Serra do Japi, Brazil, we find that the more social species, which form larger, longer-lived colonies, tend to live inside the forest, where sturdier, longer lasting vegetation is likely to offer better support for their nests. The less social species, which form single-family groups, in contrast, tend to occur on the forest edge where the vegetation is less robust. Within these two microhabitats, species with longer-lived colonies tend to occupy the potentially more stable positions closer to the core of the plants, while those with smaller and shorter-lived colonies build their nests towards the branch tips. The species further separate in their use of common habitat due to differences in the timing of their reproductive season. These patterns of habitat use suggest that the degree of sociality can enable otherwise similar species to differ from one another in ways that may facilitate their co-occurrence in a shared environment, a possibility that deserves further consideration.

A 2D/3D image analysis system to track fluorescently labeled structures in rod-shaped cells: application to measure spindle pole asymmetry during mitosis.

BACKGROUND: The yeast Schizosaccharomyces pombe is frequently used as a model for studying the cell cycle. The cells are rod-shaped and divide by medial fission. The process of cell division, or cytokinesis, is controlled by a network of signaling proteins called the Septation Initiation Network (SIN); SIN proteins associate with the SPBs during nuclear division (mitosis). Some SIN proteins associate with both SPBs early in mitosis, and then display strongly asymmetric signal intensity at the SPBs in late mitosis, just before cytokinesis. This asymmetry is thought to be important for correct regulation of SIN signaling, and coordination of cytokinesis and mitosis. In order to study the dynamics of organelles or large protein complexes such as the spindle pole body (SPB), which have been labeled with a fluorescent protein tag in living cells, a number of the image analysis problems must be solved; the cell outline must be detected automatically, and the position and signal intensity associated with the structures of interest within the cell must be determined. RESULTS: We present a new 2D and 3D image analysis system that permits versatile and robust analysis of motile, fluorescently labeled structures in rod-shaped cells. We have designed an image analysis system that we have implemented as a user-friendly software package allowing the fast and robust image-analysis of large numbers of rod-shaped cells. We have developed new robust algorithms, which we combined with existing methodologies to facilitate fast and accurate analysis. Our software permits the detection and segmentation of rod-shaped cells in either static or dynamic (i.e. time lapse) multi-channel images. It enables tracking of two structures (for example SPBs) in two different image channels. For 2D or 3D static images, the locations of the structures are identified, and then intensity values are extracted together with several quantitative parameters, such as length, width, cell orientation, background fluorescence and the distance between the structures of interest. Furthermore, two kinds of kymographs of the tracked structures can be established, one representing the migration with respect to their relative position, the other representing their individual trajectories inside the cell. This software package, called "RodCellJ", allowed us to analyze a large number of S. pombe cells to understand the rules that govern SIN protein asymmetry. CONCLUSIONS: "RodCell" is freely available to the community as a package of several ImageJ plugins to simultaneously analyze the behavior of a large number of rod-shaped cells in an extensive manner. The integration of different image-processing techniques in a single package, as well as the development of novel algorithms does not only allow to speed up the analysis with respect to the usage of existing tools, but also accounts for higher accuracy. Its utility was demonstrated on both 2D and 3D static and dynamic images to study the septation initiation network of the yeast Schizosaccharomyces pombe. More generally, it can be used in any kind of biological context where fluorescent-protein labeled structures need to be analyzed in rod-shaped cells. AVAILABILITY: RodCellJ is freely available under http://bigwww.epfl.ch/algorithms.html, (after acceptance of the publication).

Temporal SNR characteristics in segmented 3D-EPI at 7T.

Three-dimensional segmented echo planar imaging (3D-EPI) is a promising approach for high-resolution functional magnetic resonance imaging, as it provides an increased signal-to-noise ratio (SNR) at similar temporal resolution to traditional multislice 2D-EPI readouts. Recently, the 3D-EPI technique has become more frequently used and it is important to better understand its implications for fMRI. In this study, the temporal SNR characteristics of 3D-EPI with varying numbers of segments are studied. It is shown that, in humans, the temporal variance increases with the number of segments used to form the EPI acquisition and that for segmented acquisitions, the maximum available temporal SNR is reduced compared to single shot acquisitions. This reduction with increased segmentation is not found in phantom data and thus likely due to physiological processes. When operating in the thermal noise dominated regime, fMRI experiments with a motor task revealed that the 3D variant outperforms the 2D-EPI in terms of temporal SNR and sensitivity to detect activated brain regions. Thus, the theoretical SNR advantage of a segmented 3D-EPI sequence for fMRI only exists in a low SNR situation. However, other advantages of 3D-EPI, such as the application of parallel imaging techniques in two dimensions and the low specific absorption rate requirements, may encourage the use of the 3D-EPI sequence for fMRI in situations with higher SNR.

Association of BAFF/BLyS overexpression and altered B cell differentiation with Sjögren's syndrome.

BAFF (BLyS, TALL-1, THANK, zTNF4) is a member of the TNF superfamily that specifically regulates B lymphocyte proliferation and survival. Mice transgenic (Tg) for BAFF develop an autoimmune condition similar to systemic lupus erythematosus. We now demonstrate that BAFF Tg mice, as they age, develop a secondary pathology reminiscent of Sjögren's syndrome (SS), which is manifested by severe sialadenitis, decreased saliva production, and destruction of submaxillary glands. In humans, SS also correlates with elevated levels of circulating BAFF, as well as a dramatic upregulation of BAFF expression in inflamed salivary glands. A likely explanation for disease in BAFF Tg mice is excessive survival signals to autoreactive B cells, possibly as they pass through a critical tolerance checkpoint while maturing in the spleen. The marginal zone (MZ) B cell compartment, one of the enlarged B cell subsets in the spleen of BAFF Tg mice, is a potential reservoir of autoreactive B cells. Interestingly, B cells with an MZ-like phenotype infiltrate the salivary glands of BAFF Tg mice, suggesting that cells of this compartment potentially participate in tissue damage in SS and possibly other autoimmune diseases. We conclude that altered B cell differentiation and tolerance induced by excess BAFF may be central to SS pathogenesis.

Does T Helper Differentiation Correlate with Resistance or Susceptibility to Infection with L. major? Some Insights From the Murine Model.

The murine model of Leishmania major infection has been an invaluable tool in understanding T helper differentiation in vivo. The initial evidence for a role of distinct CD4(+) T helper subsets in the outcome of infection was first obtained with this experimental model. The development of CD4(+) Th1 cells was associated with resolution of the lesion, control of parasite replication, and resistance to re-infection in most of the mouse strains investigated (i.e., C57BL/6). In contrast, differentiation of CD4(+) Th2 cells correlated with the development of unhealing lesions, and failure to control parasite load in a few strains (i.e., BALB/c). Since these first reports, an incredible amount of effort has been devoted to understanding the various parameters involved in the differentiation of these, and more recently discovered T helper subsets such as Th17 and T regulatory cells. The discovery of cross-talk between T helper subsets, as well as their plasticity force us to reevaluate the events driving a protective/deleterious T helper immune response following infection with L. major in mice. In this review, we describe the individual contributions of each of these CD4(+) T helper subsets following L. major inoculation, emphasizing recent advances in the field, such as the impact of different substrains of L. major on the pathogenesis of disease.

Intracellular nanomanipulation by a photonic-force microscope with real-time acquisition of a 3D stiffness matrix.

A traditional photonic-force microscope (PFM) results in huge sets of data, which requires tedious numerical analysis. In this paper, we propose instead an analog signal processor to attain real-time capabilities while retaining the richness of the traditional PFM data. Our system is devoted to intracellular measurements and is fully interactive through the use of a haptic joystick. Using our specialized analog hardware along with a dedicated algorithm, we can extract the full 3D stiffness matrix of the optical trap in real time, including the off-diagonal cross-terms. Our system is also capable of simultaneously recording data for subsequent offline analysis. This allows us to check that a good correlation exists between the classical analysis of stiffness and our real-time measurements. We monitor the PFM beads using an optical microscope. The force-feedback mechanism of the haptic joystick helps us in interactively guiding the bead inside living cells and collecting information from its (possibly anisotropic) environment. The instantaneous stiffness measurements are also displayed in real time on a graphical user interface. The whole system has been built and is operational; here we present early results that confirm the consistency of the real-time measurements with offline computations.

The differentiation of fibre- and drug type Cannabis seedlings by gas chromatography/mass spectrometry and chemometric tools

Cannabis cultivation in order to produce drugs is forbidden in Switzerland. Thus, law enforcement authorities regularly ask forensic laboratories to determinate cannabis plant's chemotype from seized material in order to ascertain that the plantation is legal or not. As required by the EU official analysis protocol the THC rate of cannabis is measured from the flowers at maturity. When laboratories are confronted to seedlings, they have to lead the plant to maturity, meaning a time consuming and costly procedure. This study investigated the discrimination of fibre type from drug type Cannabis seedlings by analysing the compounds found in their leaves and using chemometrics tools. 11 legal varieties allowed by the Swiss Federal Office for Agriculture and 13 illegal ones were greenhouse grown and analysed using a gas chromatograph interfaced with a mass spectrometer. Compounds that show high discrimination capabilities in the seedlings have been identified and a support vector machines (SVMs) analysis was used to classify the cannabis samples. The overall set of samples shows a classification rate above 99% with false positive rates less than 2%. This model allows then discrimination between fibre and drug type Cannabis at an early stage of growth. Therefore it is not necessary to wait plants' maturity to quantify their amount of THC in order to determine their chemotype. This procedure could be used for the control of legal (fibre type) and illegal (drug type) Cannabis production.

IRF6 is a mediator of Notch pro-differentiation and tumour suppressive function in keratinocytes.

While the pro-differentiation and tumour suppressive functions of Notch signalling in keratinocytes are well established, the underlying mechanisms remain poorly understood. We report here that interferon regulatory factor 6 (IRF6), an IRF family member with an essential role in epidermal development, is induced in differentiation through a Notch-dependent mechanism and is a primary Notch target in keratinocytes and keratinocyte-derived SCC cells. Increased IRF6 expression contributes to the impact of Notch activation on growth/differentiation-related genes, while it is not required for induction of 'canonical' Notch targets like p21(WAF1/Cip1), Hes1 and Hey1. Down-modulation of IRF6 counteracts differentiation of primary human keratinocytes in vitro and in vivo, promoting ras-induced tumour formation. The clinical relevance of these findings is illustrated by the strikingly opposite pattern of expression of Notch1 and IRF6 versus epidermal growth factor receptor in a cohort of clinical SCCs, as a function of their grade of differentiation. Thus, IRF6 is a primary Notch target in keratinocytes, which contributes to the role of this pathway in differentiation and tumour suppression.

Magnetization transfer-based 3D visualization of foot peripheral nerves.

PURPOSE: To investigate magnetization transfer (MT) effects as a new source of contrast for imaging and tracking of peripheral foot nerves. MATERIALS AND METHODS: Two sets of 3D spoiled gradient-echo images acquired with and without a saturation pulse were used to generate MT ratio (MTR) maps of 260 μm in-plane resolution for eight volunteers at 3T. Scan parameters were adjusted to minimize signal loss due to T2 dephasing, and a dedicated coil was used to improve the inherently low signal-to-noise ratio of small voxels. Resulting MTR values in foot nerves were compared with those in surrounding muscle tissue. RESULTS: Average MTR values for muscle (45.5 ± 1.4%) and nerve (21.4 ± 3.1%) were significantly different (P < 0.0001). In general, the difference in MTR values was sufficiently large to allow for intensity-based segmentation and tracking of foot nerves in individual subjects. This procedure was termed MT-based 3D visualization. CONCLUSION: The MTR serves as a new source of contrast for imaging of peripheral foot nerves and provides a means for high spatial resolution tracking of these structures. The proposed methodology is directly applicable on standard clinical MR scanners and could be applied to systemic pathologies, such as diabetes.

Restricted gene flow and subpopulation differentiation in Silene dioica

The size of breeding units and the hierarchical population structure of the dioecious perennial herb Silene dioica were investigated on four closely situated island populations in the Skeppsvik Archipelago in northern Sweden. F-statistics analyses of nine polymorphic allozyme loci revealed that plants on single islands are divided into many small breeding units, between 0.2 m2 and 6 m2. Hierarchical analyses showed that levels of differentiation among subpopulations within islands (FPL=0.080) were about twice as high as among islands (FLT=0.048). These results are discussed in the light of what is known about pollen and seed movement in the archipelago.

Cyclic AMP induces differentiation in vitro of human melanoma cells.

Treating human melanoma lines with dibutyryl adenosine 3':5'-cyclic monophosphate (dbc AMP) resulted in morphologic changes associated with the altered expression of cell surface antigens. After treatment, cells developed long cellular projections characteristic of mature melanocytes and showed the presence of an increased number of Stage II premelanosomes. In addition, induction of melanin synthesis, detected as brown perinuclear pigmentation, was observed. The AMP further drastically reduced the growth rate of the five melanoma cell lines that were tested. The influence of dbc AMP was completely reversible 3 days after the agent was removed from the culture medium. The antigenic phenotype of the melanoma lines was compared before and after dbc AMP treatment. This was done with four monoclonal antibodies directed against major histocompatibility complex (MHC) Class I and II antigens and 11 monoclonal antibodies defining eight different melanoma-associated antigenic systems. Treatment with dbc AMP reduced the expression of human leukocyte antigen (HLA)-ABC antigens and beta-2-microglobulin in five of five melanoma lines. In the two HLA-DR-positive cell lines dbc AMP reduced the expression of this antigen in one line and enhanced it in the other. No induction of HLA-DR or HLA-DC antigens was observed in the Class II negative cell lines. Furthermore, dbc-AMP modulated the expression of the majority of the melanoma antigenic systems tested. The expression of a 90-kilodalton (KD) antigen, which has been found to be upregulated by interferon-gamma, was markedly decreased in all the five cell lines. A similar decrease in the expression of the high molecular weight proteoglycan-associated antigen (220-240 KD) was observed. The reduced expression of Class I and II MHC antigens as well as the altered expression of the melanoma-associated antigens studied were shown to be reversible after dbc AMP was removed. Our results collectively show that the monoclonal antibody-defined melanoma-associated molecules are linked to differentiation. They could provide useful tools for monitoring the maturation of melanomas in vivo induced by chemical agents or natural components favoring differentiation.

Dispensable role of myeloid differentiation primary response gene 88 (MyD88) and MyD88-dependent toll-like receptors (TLRs) in a murine model of osteoarthritis.

OBJECTIVES: The aim of our study was to evaluate the role of cell-membrane expressed TLRs and the signaling molecule MyD88 in a murine model of OA induced by knee menisectomy (surgical partial removal of the medial meniscus [MNX]). METHODS: OA was induced in 8-10weeks old C57Bl/6 wild-type (WT) female (n=7) mice and in knockout (KO) TLR-1 (n=7), -2 (n=8), -4 (n=9) -6 (n=5), MyD88 (n=8) mice by medial menisectomy, using the sham-operated contralateral knee as a control. Cartilage destruction and synovial inflammation were evaluated by knee joint histology using the OARSI scoring method. Apoptotic chondrocytes and cartilage metabolism (collagen II synthesis and MMP-mediated aggrecan degradation) were analyzed using immunohistochemistry. RESULTS: Operated knees exhibited OA features at 8weeks post-surgery compared to sham-operated ones. In menisectomized TLR-1, -2, -4, and -6 deficient mice, cartilage lesions, synovial inflammation and cartilage metabolism were similar to that in operated WT mice. Accordingly, using the same approach, we found no significant protection in MyD88-deficient mice in terms of OA progression as compared to WT littermates. CONCLUSIONS: Deficiency of TLRs or their signalling molecule MyD88 did not impact on the severity of experimental OA. Our results demonstrate that MyD88-dependent TLRs are not involved in this murine OA model. Moreover, the dispensable role of MyD88, which is also an adaptor for IL-1 receptor signaling, suggests that IL-1 is not a key mediator in the development of OA. This latter hypothesis is strengthened by the lack of efficiency of IL-1β antagonist in the treatment of OA.