Knowledge-based gene expression classification via matrix factorization.

MOTIVATION: Modern machine learning methods based on matrix decomposition techniques, like independent component analysis (ICA) or non-negative matrix factorization (NMF), provide new and efficient analysis tools which are currently explored to analyze gene expression profiles. These exploratory feature extraction techniques yield expression modes (ICA) or metagenes (NMF). These extracted features are considered indicative of underlying regulatory processes. They can as well be applied to the classification of gene expression datasets by grouping samples into different categories for diagnostic purposes or group genes into functional categories for further investigation of related metabolic pathways and regulatory networks. RESULTS: In this study we focus on unsupervised matrix factorization techniques and apply ICA and sparse NMF to microarray datasets. The latter monitor the gene expression levels of human peripheral blood cells during differentiation from monocytes to macrophages. We show that these tools are able to identify relevant signatures in the deduced component matrices and extract informative sets of marker genes from these gene expression profiles. The methods rely on the joint discriminative power of a set of marker genes rather than on single marker genes. With these sets of marker genes, corroborated by leave-one-out or random forest cross-validation, the datasets could easily be classified into related diagnostic categories. The latter correspond to either monocytes versus macrophages or healthy vs Niemann Pick C disease patients.

Unspecific long-term potentiation can evoke functional segregation in a model of area 17.

It has been shown recently in rat hippocampus that the synapse specificity of Hebbian long-term potentiation breaks down at short distances (< 100 microm). Using a neural network model we show that this unspecific component of long-term potentiation can be responsible for the robust formation and maintainance of cortical organization during activity-driven development. When the model is applied to the formation of orientation and ocular dominance in visual cortex, addition of an unspecific component to standard Hebbian learning, in combination with a tendency of left-eye and right-eye driven synapses to initially group together on the postsynaptic neuron, induces the simultaneous emergence and stabilization of ocular dominance and of segregated, oriented ON/OFF subfields. Since standard Hebbian learning cannot account for the simultaneous stabilization of both structures, unspecific LTP thus induces a qualitatively new behaviour. Since unspecific LTP only acts between synapses which are locally clustered in space, our results imply that details of the local grouping of synapses on the dendritic arbors of postsynaptic cells can considerably influence the formation of the cortical functional organization at the systems level.

Correction methods for three-dimensional reconstructions from confocal images: I. Tissue shrinking and axial scaling.

We show here, using locust wholemount ganglia as an example, that scaling artifacts in three-dimensional reconstructions from confocal microscopic images due to refractive index mismatch in the light path and tissue shrinking, can account for dramatic errors in measurements of morphometric values. Refractive index mismatch leads to considerable alteration of the axial dimension, and true dimensions must be restored by rescaling the Z-axis of the image stack. The appropriate scaling factor depends on the refractive indices of the media in the light path and the numerical aperture of the objective used and can be determined by numerical simulations, as we show here. In addition, different histochemical procedures were tested in regard to their effect on tissue dimensions. Reconstructions of scans at different stages of these protocols show that shrinking can be avoided prior to clearing when dehydrating ethanol series are carefully applied. Fixation and mismatching buffer osmolarity have no effect. We demonstrate procedures to reduce artifacts during mounting and clearing in methyl salicylate, such that only isometric shrinkage occurs, which can easily be corrected by rescaling the image dimensions. Glycerol-based clearing agents produced severe anisometric and nonlinear shrinkage and we could not find a way to overcome this.

A novel method for measuring saccade profiles using the scanning laser ophthalmoscope.

Most existing techniques for accurately measuring angular eye position vs time during a saccade (the saccade profile) need either contact to the eye or are restricted in time resolution. In this paper we introduce a new noninvasive method, with high spatial and temporal resolution, for determining saccade profiles using a scanning laser ophthalmoscope (SLO). This method uses the fact that images of a moving object taken with the SLO are not blurred (as are images from video cameras) but show a tilt with respect to images of the same stationary object. A mathematical framework is given that allows determination of a saccade profile from a restricted number of consecutive SLO video fields of the fundus during a saccade. The angular resolution obtained by this method is below 0.1 deg, and the maximum time resolution near 1 msec. Measured saccade profiles could be well approximated by a gamma function, the first derivative of which yields the saccade velocity profile. Measurements of peak saccade velocity as a function of saccade amplitudes (main sequence) using our method show good agreement with results obtained by other authors.

Blind signal separation from optical imaging recordings with extended spatial decorrelation.

Optical imaging is the video recording of two-dimensional patterns of changes in light reflectance from cortical tissue evoked by stimulation. We derived a method, extended spatial decorrelation (ESD), that uses second-order statistics in space for separating the intrinsic signals into the stimulus related components and the nonspecific variations. The performance of ESD on model data is compared to independent component analysis algorithms using statistics of fourth and higher order. Robustness against sensor noise is scored. When applied to optical images, ESD separates the stimulus specific signal well from biological noise and artifacts.

Evaluation of sensitivity for wild type and mutant forms of hepatitis B surface antigen by four commercial HBsAg assays.

Hepatitis B surface antigen is one of the most important serological markers used to diagnose an HBV infection. Improvements in HBsAg assay sensitivity have been achieved constantly over the years since introduction of the first commercial assays. It is generally assumed that diagnostic assay sensitivity includes the ability to detect wild type and viral variants at the same level. Thus it would be expected, as newer HBsAg assays are developed, that viral mutation would be considered in assay design and that assay sensitivity would be equivalent for wild type as well as mutant forms. Two newly launched HBsAg assays (Bayer ADVIA Centaur and Ortho VITROS ECi) were compared to two established HBsAg assays (Abbott AxSYM and Roche Elecsys) in order to test the assumption that assay sensitivity for variants is equivalent to wild type HBsAg. The four assays were challenged with a standard HBsAg sensitivity panel of both ad and ay subtypes as well as a 13 member mutant panel comprised of both recombinant and native HBsAg members. Results demonstrate that the analytical sensitivity for wild type HBsAg is comparable for all assays tested. In contrast, significant differences were observed for detection of mutants. AxSYM HBsAg detected all mutant samples while all other asssays missed 10 out of 13 samples tested. It is noteworthy that the most frequently reported HBsAg mutation, G145 R, remained undetected in three of the assays tested. It is discussed whether the reduced sensitivity for mutants of the most recent assays represents a new risk for the diagnosis of HBV infection.

Acute Sarcocystis falcatula-like infection in a carmine bee-eater (Merops nubicus) and immunohistochemical cross reactivity between Sarcocystis falcatula and Sarcocystis neurona.

An unidentified Sarcocystis falcatula-like infection was diagnosed in a captive bee-eater (Merops nubicus) in a zoo in Florida. The bird died suddenly, probably due to protozoa-associated pneumonia. Protozoal schizonts were found in lungs and heart, and immature sarcocysts were seen in skeletal muscles. Ultrastructurally, schizonts were located in capillary endothelium and merozoites lacked rhoptries, consistent with the structure of Sarcocystis species. Sarcocysts were immature, microscopic, and contained only metrocytes. The sarcocyst wall had finger-like villar protrusions that were up to 0.7 microm long and up to 0.2 microm wide. The villar protrusions lacked microtubules, characteristically seen in sarcocysts of S. falcatula. Antigenically, parasites in lungs and muscles of the bee-eater reacted with a varying intensity with polyclonal rabbit antisera to S. falcatula and Sarcocystis neurona. Results indicated that sarcocysts in the bee-eater were morphologically different from the reported structure for sarcocysts of other S. falcatula infections.

Rheological properties of blood as assessed with a newly designed oscillating viscometer.

A newly designed type of oscillating viscometer was tested for blood viscosity measurements. The viscometer consists of a probe (either a tube or a rod) oscillating at a resonance frequency with amplitudes in the micro- and nanometer range. The torsional oscillations are dampened by fluids flowing through the tube or surrounding the rod. The degree of damping depends on the viscosity of the fluid, which allows to measure viscosity. Data obtained with these instruments were compared with those obtained with a conventional Couette viscometer. An increase of erythrocyte aggregation by the addition of dextran 70 in vitro led to the expected increase of viscosity in the Couette viscometer; in the oscillating tube viscometer, however, it remained unchanged, which may be explained by a decreased erythrocyte concentration near the tube wall due to increased aggregation and flow of erythrocytes in the tube center. In ex vivo experiments on blood flowing without anticoagulant directly through the tube viscometer an inverse correlation between viscosity and fibrinogen concentration was found. This is in contrast to actual knowledge and may indicate that high fibrinogen levels have a beneficial rheological effect at the tube or vessel wall. Our data suggest that the new oscillating tube viscometer is an interesting tool, which may contribute to a more comprehensive understanding of blood flow.

Femoral artery pseudoaneurysm in a monkey.

Pseudoaneurysm formation as a complication of routine blood collection was diagnosed in a monkey. Damage to the femoral artery resulted in hematoma formation with secondary organization, encapsulation, and vascular communication. Progressive lameness and muscular atrophy were the primary clinical signs. Surgical correction of the artery defect helped resolve the monkey's lameness and muscle atrophy.

Epizootic of Mycobacterium bovis in a zoologic park.

An epizootic of Mycobacterium bovis in a zoologic park resulted in the death of 4 southern white rhinoceroses and 2 colobus monkeys. Zoo personnel were detected that had positive intradermal tuberculin skin test results after exposure to mycobacterial-infected animals. On the basis of DNA fingerprinting, all 3 mycobacterial isolates (from 1 rhinoceros and 2 monkeys) were determined to be genetically similar and probably originated from the same source. The 3 animals (1 rhinoceros and 2 colobus monkeys) that had confirmed infections lived in separate, but adjacent, areas. Aerosolization of bacteria during routine cleaning was believed to have contributed to the unusual distance between infected animals. Tuberculosis has reemerged as a major disease problem in human and veterinary medicine.

A neural network model for the self-organization of cortical grating cells.

A neural network model with incremental Hebbian learning of afferent and lateral synaptic couplings is proposed,which simulates the activity-dependent self-organization of grating cells in upper layers of striate cortex. These cells, found in areas V1 and V2 of the visual cortex of monkeys, respond vigorously and exclusively to bar gratings of a preferred orientation and periodicity. Response behavior to varying contrast and to an increasing number of bars in the grating show threshold and saturation effects. Their location with respect to the underlying orientation map and their nonlinear response behavior are investigated. The number of emerging grating cells is controlled in the model by the range and strength of the lateral coupling structure.

Fish and amphibian anesthesia.

This article is designed to provide the veterinary practitioner with some basic information on how to anesthetize fish and amphibians. General guidelines regarding working with these aquatic animals are covered. Specific topics include methods of anesthetic delivery, anesthetic monitoring, and recovery. Various anesthetic regimes and dosages are presented both in the text and in table form.

Immobilization of babirusa (Babyrousa babyrussa) with xylazine and tiletamine/zolazepam and reversal with yohimbine and flumazenil.

Twelve babirusa (Babyrousa babyrussa) (four females/eight males) were immobilized 30 times during a 4-yr interval. Significantly higher premedication and immobilizing doses were needed for females than for males (P < 0.05). An i.m. preanesthetic xylazine dose of 1.88 +/- 0.37 mg/kg (range = 1.20-2.12 mg/kg) was used for females and 1.22 +/- 0.16 mg/kg (range = 0.82-1.43 mg/kg) for males. After xylazine, the animals were induced with i.m. tiletamine/zolazepam; females received 2.20 +/- 0.47 mg/kg (range = 1.78-3.33 mg/kg) and males received 1.71 +/- 0.34 mg/kg (range = 1.08-2.05 mg/kg). Anesthesia was reversed with yohimbine (0.14 +/- 0.03 mg/kg; range = 0.07-0.20 mg/kg) and flumazenil (1 mg flumazenil/20 mg zolazepam) either i.m. or i.v. This anesthetic combination produced smooth induction, good relaxation, and sufficient immobilization to perform routine diagnostic and therapeutic procedures (venipuncture, hoof and tusk trims, transportation, radiographs, ultrasound examination, weight determinations, and skin biopsies). Supplemental ketamine HCl or isoflurane was administered to two animals to effectively deepen or prolong the anesthetic plane, with no resultant adverse effects.

Simulation of scanning laser techniques for optical imaging of blood-related intrinsic signals.

Optical imaging of intrinsic signals detects neural activation patterns by taking video images of the local activity-related changes in the light intensity reflected from neural tissue (intrinsic signals). At red light (605 nm), these signals are caused mainly by local variations of the tissue absorption following deoxygenation of blood. We characterize the image generation process during optical imaging by Monte Carlo simulations of light propagation through a homogeneous model tissue equipped with a local absorber. Conventional video imaging and scanning laser imaging are compared. We find that, compared with video imaging, scanning laser techniques drastically increase both the contrast and the lateral resolution of optical recordings. Also, the maximum depth up to which the signals can be detected is increased by roughly a factor of 2 when scanning laser optical imaging is used. Further, the radial profile of the diffuse-reflectance pattern for each pixel is subject to changes that correlate with the depth of the absorber within the tissue. We suggest a detection geometry for the online measurement of these radial profiles that can be realized by modifying a standard scanning laser ophthalmoscope.

Azathioprine hypersensitivity mimicking Goodpasture's syndrome.

Side effects due to azathioprine (the nitroimidazole derivative of 6-mercaptopurine) can be classified as toxic (myelosuppression, hepatotoxicity) and idiosyncratic (fever, rigors, arthralgias, pneumonitis, and gastrointestinal symptoms). While the toxic effects are due to 6-mercaptopurine, the hypersensitivity reactions are believed to be caused by the nitroimidazole moiety. A 21-year-old male patient developed end-stage renal failure due to antiglomerular basement membrane (AGBM) disease (rapidly progressive glomerulonephritis with linear immunoglobulin G deposits and positive circulating AGBM antibodies). The patient became dependent on continuous ambulatory peritoneal dialysis and, later, hemodialysis, and received two renal allografts at the ages of 23 and 27 years. He received three courses of azathioprine treatment: one course for AGBM glomerulonephritis and two courses for rejection episodes. Each course was followed within 4 to 7 days by symptoms compatible with Goodpasture's syndrome, ie, high fever, rigors, arthralgias, diarrhea, myalgias, and pulmonary infiltrates with hemoptysis. All signs and symptoms always resolved completely on discontinuation of azathioprine. During the treatment for rejections, AGBM antibodies were not elevated, and during one episode AGBM disease in the lung (Goodpasture's syndrome) was excluded by open lung biopsy. Treatment of a subsequent rejection episode with 6-mercaptopurine was well tolerated. We conclude that azathioprine hypersensitivity can mimic the pulmonary manifestations of Goodpasture's syndrome. Hypersensitivity probably is due to the nitroimidazole moiety of azathioprine. Thus, differential diagnosis of Goodpasture's syndrome (and probably of any "pulmonary renal syndrome") should include azathioprine hypersensitivity.

[Long-term follow-up of Crohn disease]. / Langzeitverlauf des Morbus Crohn.

184 patients diagnosed with Crohn's disease were observed for an average of 5 years and evaluated for clinical pattern and course of disease. The male/female ratio was almost equal (90 m. vs. 94 f.). The peak of disease manifestation was between years 18-32. Familial clustering of inflammatory bowel disease was observed in 2.2%. The pattern of anatomic involvement was ileocolic in 53%, colon in 24%, small bowel in 20% and other locations in 3%. 34% of patients were afflicted with at least one extraintestinal manifestation, 34% developed fistulae. Fistula formation was seen more often in colon involvement than small bowel involvement (69% vs. 32%). 48% of the patients were operated at least once. 3 patients died during the observation period, 2 of them with sepsis. Evolution of carcinoma was not observed. Some of our results are in accordance with the literature, some distinct observations (e.g. low familial clustering) underline the theory of a multifactorial pathogenesis in Crohn's disease.

A mean-field model for orientation tuning, contrast saturation, and contextual effects in the primary visual cortex.

Orientation-selective cells in the primary visual cortex of monkeys and cats are often characterized by an orientation-tuning width that is invariant under stimulus contrast. At the same time their contrast response function saturates or even super-saturates for high values of contrast. When two bar stimuli are presented within their classical receptive field, the neuronal response decreases with the intersection angle. When two stimuli are presented inside and outside the classical receptive field, the response of the cell increases with the intersection angle. Both cats and monkeys show iso-orientation suppression, which has sometimes been reported to be combined with cross-orientation facilitation. This property has previously been described as sensitivity to orientation contrast. We address the emergence of these effects with a model that describes the processing of geniculocortical signals through cortical circuitry. We hypothesize that short intracortical fibers mediate the classical receptive field effects, whereas long-range collaterals evoke contextual effects such as sensitivity to orientation contrast. We model this situation by setting up a mean-field description of two neighboring cortical hypercolumns, which can process a nonoverlapping center and a (nonclassical) surround stimulus. Both hypercolumns interact via idealized long-range connections. For an isolated model hypercolumn, we find that either contrast saturation or contrast-invariant orientation tuning emerges, depending on the strength of the lateral excitation. There is no parameter regime, however, where both phenomena emerge simultaneously. In the regime where contrast saturation is found, the model also correctly reproduces suppression due to a second, cross-oriented grid within the classical receptive field. If two model hypercolumns are mutually coupled by long-range connections that are iso-orientation specific, nonclassical surround stimuli show either suppression or facilitation for all surround orientations. Sensitivity to orientation contrast is not observed. This property requires excitatory-to-excitatory long-range couplings that are less orientation specific than those targeting inhibitory neurons.

Analysis of calcium imaging signals from the honeybee brain by nonlinear models.

Recent Ca(2+)-imaging studies on the antennal lobe of the honeybee (Apis mellifera) have shown that olfactory stimuli evoke complex spatiotemporal changes of the intracellular Ca(2+) concentration, in which stimulus-dependent subsets of glomeruli are highlighted. In this work we use nonlinear models for the quantitative identification of the spatial and temporal properties of the Ca(2+)-dependent fluorescence signal. This technique describes time series of the Ca(2+) signal as a superposition of biophysically motivated model functions for photobleaching and Ca(2+) dynamics and provides optimal estimates of their amplitudes (signal strengths) and time constants together with error measures. Using this method, we can reliably identify two different stimulus-dependent signal components. Their delays and rise times, delta(c1) = (0.4 +/- 0.3) s, tau(c1) = (3.8 +/- 1.2) s for the fast component and delta(c2) = (2.4 +/- 0.6) s, tau(c2) = (10.3 +/- 3.2) s for the slow component, are constant over space and across different odors and animals. In chronological experiments, the amplitude of the fast (slow) component often decreases (increases) with time. The pattern of the Ca(2+) dynamics in space and time can be reliably described as a superposition of only two spatiotemporally separable patterns based on the fast and slow components. However, the distributions of both components over space turn out to differ from each other, and more work has to be done in order to specify their relationship with neuronal activity.

Rhinoceros' rhinorrhea: cause of an outbreak of infection due to airborne Mycobacterium bovis in zookeepers.

Seven of 24 zookeepers exposed to a Southern white rhinoceros infected with Mycobacterium bovis were presumably infected via aerosols generated in the cleaning of the barn for the rhinoceros. All demonstrated conversion by the intermediate-strength purified-protein-derivative skin test, but none had clinical illness. In certain occupational settings like zoos and abattoirs, exposure to M. bovis may be an occupational hazard, and routine periodic tuberculin screening should be performed.

A neural network model for the emergence of grating cells.

A correlation-based learning (CBL) neural network model is proposed, which simulates the emergence of grating cells as well as some of their response characteristics to periodic pattern stimuli. These cells, found in areas V1 and V2 of the visual cortex of monkeys, respond vigorously and exclusively to bar gratings of a preferred orientation and periodicity. Their non-linear behaviour differentiates grating cells from other orientation-selective cells, which show linear spatial frequency filtering.