Unsupervised clustering of fMRI and MRI time series

Unsupervised clustering represents a powerful technique for self-organized segmentation of biomedical image time series data describing groups of pixels exhibiting similar properties of local signal dynamics. The theoretical background is presented in the beginning, followed by several medical applications demonstrating the flexibility and conceptual power of these techniques. These applications range from functional MRI data analysis to dynamic contrast-enhanced perfusion MRI and breast MRI. For fMRI, these methods can be employed to identify and separate time courses of interest, along with their associated spatial patterns. When applied to dynamic perfusion MRI, they identify groups of voxels associated with time courses that are clinically informative and straightforward to interpret. In breast MRI, a segmentation of the lesion is achieved and in addition a subclassification is obtained within the lesion with regard to regions characterized by different MRI signal time courses. In the present paper, we conclude that unsupervised clustering techniques provide a robust method for blind analysis of time series image data in the important and current field of functional and dynamic MRI.     

Tissue-growth-based synthetic tree generation and perfusion simulation

Biological tissues receive oxygen and nutrients from blood vessels by developing an indispensable supply and demand relationship with the blood vessels. We implemented a synthetic tree generation algorithm by considering the interactions between the tissues and blood vessels. We first segment major arteries using medical image data and synthetic trees are generated originating from these segmented arteries. They grow into extensive networks of small vessels to fill the supplied tissues and satisfy the metabolic demand of them. Further, the algorithm is optimized to be executed in parallel without affecting the generated tree volumes. The generated vascular trees are used to simulate blood perfusion in the tissues by performing multiscale blood flow simulations. One-dimensional blood flow equations were used to solve for blood flow and pressure in the generated vascular trees and Darcy flow equations were solved for blood perfusion in the tissues using a porous model assumption. Both equations are coupled at terminal segments explicitly. The proposed methods were applied to idealized models with different tree resolutions and metabolic demands for validation. The methods demonstrated that realistic synthetic trees were generated with significantly less computational expense compared to that of a constrained constructive optimization method. The methods were then applied to cerebrovascular arteries supplying a human brain and coronary arteries supplying the left and right ventricles to demonstrate the capabilities of the proposed methods. The proposed methods can be utilized to quantify tissue perfusion and predict areas prone to ischemia in patient-specific geometries.

     

Adaptive thermal modeling: a concept for measurement of local blood perfusion in heated tissues

A new Adaptive Thermal Modeling (ATM) method for the measurement of local tissue blood perfusion rate is introduced. The method is based on a two-phase numerical technique. The first phase includes a fast, finite difference scheme for solution of the transient temperature field. The second phase involves iterative corrections of the perfusion until the modeled temperatures coincide with those measured by the temperature sensors. The results obtained from computer generated "data", as well as from laboratory experiments demonstrate the potential capability of the ATM method to continuously measure local perfusion rates in heated tissues. Rigorous analysis of the technique is planned for the near future so that it can be applied to in vivo measurements of local tissue blood perfusions.     

Colorization and automated segmentation of human T2 MR brain images for characterization of soft tissues

Characterization of tissues like brain by using magnetic resonance (MR) images and colorization of the gray scale image has been reported in the literature, along with the advantages and drawbacks. Here, we present two independent methods; (i) a novel colorization method to underscore the variability in brain MR images, indicative of the underlying physical density of bio tissue, (ii) a segmentation method (both hard and soft segmentation) to characterize gray brain MR images. The segmented images are then transformed into color using the above-mentioned colorization method, yielding promising results for manual tracing. Our color transformation incorporates the voxel classification by matching the luminance of voxels of the source MR image and provided color image by measuring the distance between them. The segmentation method is based on single-phase clustering for 2D and 3D image segmentation with a new auto centroid selection method, which divides the image into three distinct regions (gray matter (GM), white matter (WM), and cerebrospinal fluid (CSF) using prior anatomical knowledge). Results have been successfully validated on human T2-weighted (T2) brain MR images. The proposed method can be potentially applied to gray-scale images from other imaging modalities, in bringing out additional diagnostic tissue information contained in the colorized image processing approach as described.     

Cerebrovascular casting of the adult mouse for 3D imaging and morphological analysis

Vascular imaging is crucial in the clinical diagnosis and management of cerebrovascular diseases, such as brain arteriovenous malformations (BAVMs). Animal models are necessary for studying the etiopathology and potential therapies of cerebrovascular diseases. Imaging the vasculature in large animals is relatively easy. However, developing vessel imaging methods of murine brain disease models is desirable due to the cost and availability of genetically-modified mouse lines. Imaging the murine cerebral vascular tree is a challenge. In humans and larger animals, the gold standard for assessing the angioarchitecture at the macrovascular (conductance) level is x-ray catheter contrast-based angiography, a method not suited for small rodents. In this article, we present a method of cerebrovascular casting that produces a durable skeleton of the entire vascular bed, including arteries, veins, and capillaries that may be analyzed using many different modalities. Complete casting of the microvessels of the mouse cerebrovasculature can be difficult; however, these challenges are addressed in this step-by-step protocol. Through intracardial perfusion of the vascular casting material, all vessels of the body are casted. The brain can then be removed and clarified using the organic solvent methyl salicylate. Three dimensional imaging of the brain blood vessels can be visualized simply and inexpensively with any conventional brightfield microscope or dissecting microscope. The casted cerebrovasculature can also be imaged and quantified using micro-computed tomography (micro-CT)(1). In addition, after being imaged, the casted brain can be embedded in paraffin for histological analysis. The benefit of this vascular casting method as compared to other techniques is its broad adaptation to various analytic tools, including brightfield microscopic analysis, CT scanning due to the radiopaque characteristic of the material, as well as histological and immunohistochemical analysis. This efficient use of tissue can save animal usage and reduce costs. We have recently demonstrated application of this method to visualize the irregular blood vessels in a mouse model of adult BAVM at a microscopic level(2), and provide additional images of the malformed vessels imaged by micro-CT scan. Although this method has drawbacks and may not be ideal for all types of analyses, it is a simple, practical technique that can be easily learned and widely applied to vascular casting of blood vessels throughout the body.     

Walsh Hadamard Transform for Simple Linear Iterative Clustering (SLIC) Superpixel Based Spectral Clustering of Multimodal MRI Brain Tumor Segmentation

The automated brain tumor segmentation methods are challenging due to the diverse nature of tumors. Recently, the graph based spectral clustering method is utilized for brain tumor segmentation to make high-quality segmentation output. In this paper, a new Walsh Hadamard Transform (WHT) texture for superpixel based spectral clustering is proposed for segmentation of a brain tumor from multimodal MRI images. First, the selected kernels of WHT are utilized for creating texture saliency maps and it becomes the input for the Simple Linear Iterative Clustering (SLIC) algorithm, to generate more precise texture based superpixels. Then the texture superpixels become nodes in the graph of spectral clustering for segmenting brain tumors of MRI images. Finally, the original members of superpixels are recovered to represent Complete Tumor (CT), Tumor Core (TC) and Enhancing Tumor (ET) tissues. The observational results are taken out on BRATS 2015 datasets and evaluated using the Dice Score (DS), Hausdorff Distance (HD) and Volumetric Difference (VD) metrics. The proposed method produces competitive results than other existing clustering methods.     

Absent MicroRNAs in Different Tissues of Patients with Acquired Cardiomyopathy

MicroRNAs (miRNAs) can be found in a wide range of tissues and body fluids, and their specific signatures can be used to determine diseases or predict clinical courses. The miRNA profiles in biological samples (tissue, serum, peripheral blood mononuclear cells or other body fluids) differ significantly even in the same patient and therefore have their own specificity for the presented con-dition. Complex profiles of deregulated miRNAs are of high interest, whereas the importance of non-expressed miRNAs was ignored. Since miRNAs regulate gene expression rather negatively, absent miRNAs could indicate genes with unaltered expression that therefore are normally expressed in specific compartments or under specific disease situations. For the first time, non-detectable miRNAs in different tissues and body fluids from patients with different diseases (cardiomyopathies, Alzheimer’s disease, bladder cancer, and ocular cancer) were analyzed and com-pared in this study. miRNA expression data were generated by microarray or TaqMan PCR-based platforms. Lists of absent miRNAs of primarily cardiac patients (myocardium, blood cells, and serum) were clustered and analyzed for potentially involved pathways using two prediction platforms, i.e., miRNA enrichment analysis and annotation tool (miEAA) and DIANA miRPath. Extensive search in biomedical publication databases for the relevance of non-expressed miRNAs in predicted pathways revealed no evidence for their involvement in heart-related pathways as indicated by software tools, confirming proposed approach.     

Rough-Fuzzy Clustering and Unsupervised Feature Selection for Wavelet Based MR Image Segmentation

Image segmentation is an indispensable process in the visualization of human tissues, particularly during clinical analysis of brain magnetic resonance (MR) images. For many human experts, manual segmentation is a difficult and time consuming task, which makes an automated brain MR image segmentation method desirable. In this regard, this paper presents a new segmentation method for brain MR images, integrating judiciously the merits of rough-fuzzy computing and multiresolution image analysis technique. The proposed method assumes that the major brain tissues, namely, gray matter, white matter, and cerebrospinal fluid from the MR images are considered to have different textural properties. The dyadic wavelet analysis is used to extract the scale-space feature vector for each pixel, while the rough-fuzzy clustering is used to address the uncertainty problem of brain MR image segmentation. An unsupervised feature selection method is introduced, based on maximum relevance-maximum significance criterion, to select relevant and significant textural features for segmentation problem, while the mathematical morphology based skull stripping preprocessing step is proposed to remove the non-cerebral tissues like skull. The performance of the proposed method, along with a comparison with related approaches, is demonstrated on a set of synthetic and real brain MR images using standard validity indices.     

Blue laser light increases perfusion of a skin flap via release of nitric oxide from hemoglobin

It has recently been shown that nitrosyl complexes of hemoglobin (NO-Hb) are sensitive to low-level blue laser irradiation, suggesting that laser irradiation can facilitate the release of biologically active nitric oxide (NO), which can affect tissue perfusion. The aim of this study was to evaluate the therapeutic value of blue laser irradiation for local tissue perfusion after surgical intervention. Blood was withdrawn from a rat, exposed to NO and infused back to the same rat or used for in vitro experiments. In vitro, an increase of NO-Hb levels (electron paramagnetic resonance spectroscopy) up to 15 microM in rat blood did not result in the release of detectable amounts of NO (NO selective electrode). Blue laser irradiation of NO-Hb in blood caused decomposition of NO-Hb complexes and release of free NO. Systemic infusion of NO-Hb in rats affected neither systemic circulation (mean arterial pressure) nor local tissue perfusion (Doppler blood flow imaging system). In contrast, a clear enhancement of local tissue perfusion was observed in epigastric flap when elevated NO-Hb levels in blood were combined with local He-Cd laser irradiation focused on the left epigastric artery. The enhancement of regional tissue perfusion was not accompanied by any detectable changes in systemic circulation. This study demonstrates that blue laser irradiation improves local tissue perfusion in a controlled manner stimulating NO release from NO-Hb complexes.     

Genome-wide profiling of gene expression in 29 normal human tissues with a cDNA microarray.

We have performed a comprehensive analysis of the expression profiles in 25 adult and 4 fetal human tissues by means of a cDNA microarray consisting of 23,040 human genes. This study revealed a number of genes that were expressed specifically in each of those tissues. Among the 29 tissues examined, 4,080 genes were highly expressed (at least a five-fold expression ratio) in one or only a few tissues and 1,163 of those were expressed exclusively (more than a ten-fold higher expression ratio) in a particular tissue. Expression of some of the genes in the latter category was confirmed by northern analysis. A hierarchical clustering analysis of gene-expression profiles in nerve tissues (adult brain, fetal brain, and spinal cord), lymphoid tissues (bone marrow, thymus, spleen, and lymph node), muscle tissues (heart and skeletal muscle), or adipose tissues (mesenteric adipose and mammary gland) identified a set of genes that were commonly expressed among related tissues. These data should provide useful information for medical research, especially for efforts to identify tissue-specific molecules as potential targets of novel drugs to treat human diseases.     

血脑屏障与脑血管疾病的相关研究

血脑屏障（blood brain barrier,BBB）的主要结构包括：脑毛细血管内皮细胞及其间的紧密连接（tight junction,TJ）、基底膜、基 底膜下星型胶质细胞终足。血脑屏障是存在于血液和脑组织之间的一层屏障系统,在许多大脑疾患的病理过程中,BBB 的破坏导 致通透性增高都是不可避免的一个环节。BBB是保证中枢神经系统的正常生理功能的重要屏障系统。目前已有大量关于血脑屏 障通透性在脑血管疾病中的变化研究。本文分别从血脑屏障的结构和功能,药物通过血脑屏障的方法和功能,脑缺血损伤、阿尔 茨海默病、帕金森病和多发性硬化症等不同的脑病变与血脑屏障通透性的变化及中医药应用等方面做一综述。有针对性地对 BBB和大脑疾病进行进一步的研究与探索,将会为临床治疗相关疾病带来新的视角与机遇。     

Targeting Therapeutics Across the Blood Brain Barrier (BBB), Prerequisite Towards Thrombolytic Therapy for Cerebrovascular Disorders—an Overview and Advancements

Cerebral tissues possess highly selective and dynamic protection known as blood brain barrier (BBB) that regulates brain homeostasis and provides protection against invading pathogens and various chemicals including drug molecules. Such natural protection strictly monitors entry of drug molecules often required for the management of several diseases and disorders including cerebral vascular and neurological disorders. However, in recent times, the ischemic cerebrovascular disease and clinical manifestation of acute arterial thrombosis are the most common causes of mortality and morbidity worldwide. The management of cerebral Ischemia requires immediate infusion of external thrombolytic into systemic circulation and must cross the blood brain barrier. The major challenge with available thrombolytic is their poor affinity towards the blood brain barrier and cerebral tissue subsequently. In the clinical practice, a high dose of thrombolytic often prescribed to deliver drugs across the blood brain barrier which results in drug dependent toxicity leading to damage of neuronal tissues. In recent times, more emphasis was given to utilize blood brain barrier transport mechanism to deliver drugs in neuronal tissue. The blood brain barrier expresses a series of receptor on membrane became an ideal target for selective drug delivery. In this review, the author has given more emphasis molecular biology of receptor on blood brain barrier and their potential as a carrier for drug molecules to cerebral tissues. Further, the use of nanoscale design and real-time monitoring for developed therapeutic to encounter drug dependent toxicity has been reviewed in this study.KEY WORDS: blood brain barrier (BBB), cerebral ischemic disorders, drug delivery, earthworm protease, neurodegenerative disorder, thrombolytic     

Neurovascular and Cognitive failure in Alzheimer’s Disease: Benefits of Cardiovascular Therapy

Alzheimer’s disease (AD) is a multifactorial and multifaceted disease for which we currently have very little to offer since there is no curative therapy, with only limited disease-modifying drugs. Recent studies in AD mouse models that recapitulate the amyloid-β (Aβ) pathology converge to demonstrate that it is possible to salvage cerebrovascular function with a variety of drugs and, particularly, therapies used to treat cardiovascular diseases such as hypercholesterolemia and hypertension. These drugs can reestablish dilatory function mediated by various endothelial and smooth muscle ion channels as well as nitric oxide availability, benefits that result in normalized brain perfusion. These cerebrovascular benefits would favor brain perfusion, which may help maintain neuronal function and, possibly, delay cognitive failure. However, restoring cerebrovascular function in AD mouse models was not necessarily accompanied by rescue of cognitive deficits related to spatial learning and memory. The results with cardiovascular therapies rather suggest that drugs originally designed to treat cardiovascular diseases that concurrently restore cerebrovascular and cognitive function do so through their pleiotropic effects. Specifically, recent findings suggest that these drugs act directly on brain cells and neuronal pathways involved in memory formation, hence, working simultaneously albeit independently on neuronal and vascular targets. These findings may help select medications for patients with cardiovascular diseases at risk of developing AD with increasing age. Further, they may identify molecular targets for recovering memory pathways that bear potential for new therapeutic avenues.     

In Situ Detection of Bacteria within Paraffin-embedded Tissues Using a Digoxin-labeled DNA Probe Targeting 16S rRNA

The presence of bacteria within the pocket epithelium and underlying connective tissue in gingival biopsies from patients with periodontitis has been reported using various methods, including electron microscopy, immunohistochemistry or immunofluorescence using bacteria-specific antibodies, and fluorescent in situ hybridization (FISH) using a fluorescence-labeled oligonucleotide probe. Nevertheless, these methods are not widely used due to technical limitation or difficulties. Here a method to localize bacteria within paraffin-embedded tissues using DIG-labeled DNA probes has been introduced. The paraffin-embedded tissues are the most common form of biopsy tissues available from pathology banks. Bacteria can be detected either in a species-specific or universal manner. Bacterial signals are detected as either discrete forms (coccus, rod, fusiform, and hairy form) of bacteria or dispersed forms. The technique allows other histological information to be obtained: the epithelia, connective tissue, inflammatory infiltrates, and blood vessels are well distinguished. This method can be used to study the role of bacteria in various diseases, such as periodontitis, cancers, and inflammatory immune diseases.     

A numerical model of the fracture healing process that describes tissue development and revascularisation

A dynamic model was developed to simulate complex interactions of mechanical stability, revascularisation and tissue differentiation in secondary fracture healing. Unlike previous models, blood perfusion was included as a spatio-temporal state variable to simulate the revascularisation process. A 2D, axisymmetrical finite element model described fracture callus mechanics. Fuzzy logic rules described the following biological processes: angiogenesis, intramembranous ossification, chondrogenesis, cartilage calcification and endochondral ossification, all of which depended on local strain state and local blood perfusion. In order to evaluate how the predicted revascularisation depended on the mechanical environment, we simulated two different healing cases according to two groups of transverse metatarsal osteotomies in sheep with different axial stability. The model predicted slower revascularisation and delayed bony bridging for the less stable case, which corresponded well to the experimental observations. A revascularisation sensitivity analysis demonstrated the potential of the model to account for different conditions regarding the blood supply.     

A numerical model of the fracture healing process that describes tissue development and revascularisation

A dynamic model was developed to simulate complex interactions of mechanical stability, revascularisation and tissue differentiation in secondary fracture healing. Unlike previous models, blood perfusion was included as a spatio-temporal state variable to simulate the revascularisation process. A 2D, axisymmetrical finite element model described fracture callus mechanics. Fuzzy logic rules described the following biological processes: angiogenesis, intramembranous ossification, chondrogenesis, cartilage calcification and endochondral ossification, all of which depended on local strain state and local blood perfusion. In order to evaluate how the predicted revascularisation depended on the mechanical environment, we simulated two different healing cases according to two groups of transverse metatarsal osteotomies in sheep with different axial stability. The model predicted slower revascularisation and delayed bony bridging for the less stable case, which corresponded well to the experimental observations. A revascularisation sensitivity analysis demonstrated the potential of the model to account for different conditions regarding the blood supply.     

Evaluation of laser-Doppler flowmetry as a measure of tissue blood flow

In this study the technique of laser-Doppler flowmetry was evaluated for the measurement of tissue blood flow by comparing laser-Doppler flow (LDF) signal in the renal cortex, gracilis muscle, and cremaster muscle of anesthetized rats to whole-organ blood flow measured with an electromagnetic flowmeter or radioactive microspheres. In vitro, LDF signal was closely correlated (r = 0.99) to changes in erythrocyte velocity generated with a rotating wheel. Although individual LDF readings varied in situ, mean LDF signal calculated from multiple readings on the tissue surface were significantly correlated (r = 0.74-0.95) with tissue blood flows measured at various perfusion pressures. However, significant differences in the slope of the LDF signal vs. blood flow relationship were observed in different tissues and with different methods of measurement in the same tissue. This study indicates that mean laser-Doppler flow signal provides a good estimate of tissue blood flow, provided a sufficient number of points is scanned. However, there appears to be no universal calibration factor for the method.     

Mechanisms of the Anti-Ischemic Effect of Angiotensin II AT( 1 ) Receptor Antagonists in the Brain

1. Circulating and locally formed Angiotensin II regulates the cerebral circulation through stimulation of AT(1) receptors located in cerebrovascular endothelial cells and in brain centers controlling cerebrovascular flow. 2. The cerebrovascular autoregulation is designed to maintain a constant blood flow to the brain, by vasodilatation when blood pressure decreases and vasoconstriction when blood pressure increases. 3. During hypertension, there is a shift in the cerebrovascular autoregulation to the right, in the direction of higher blood pressures, as a consequence of decreased cerebrovascular compliance resulting from vasoconstriction and pathological growth. In hypertension, when perfusion pressure decreases as a consequence of blockade of a cerebral artery, reduced cerebrovascular compliance results in more frequent and more severe strokes with a larger area of injured tissue. 4. There is a cerebrovascular angiotensinergic overdrive in genetically hypertensive rats, manifested as an increased expression of cerebrovascular AT(1) receptors and increased activity of the brain Angiotensin II system. Excess AT(1) receptor stimulation is a main factor in the cerebrovascular pathological growth and decreased compliance, the alteration of the cerebrovascular eNOS/iNOS ratio, and in the inflammatory reaction characteristic of cerebral blood vessels in genetic hypertension. All these factors increase vulnerability to brain ischemia and stroke. 5. Sustained blockade of AT(1) receptors with peripheral and centrally active AT(1) receptor antagonists (ARBs) reverses the cerebrovascular pathological growth and inflammation, increases cerebrovascular compliance, restores the eNOS/iNOS ratio and decreases cerebrovascular inflammation. These effects result in a reduction of the vulnerability to brain ischemia, revealed, when an experimental stroke is produced, in protection of the blood flow in the zone of penumbra and substantial reduction in neuronal injury. 6. The protection against ischemia resulting is related to inhibition of the Renin-Angiotensin System and not directly related to the decrease in blood pressure produced by these compounds. A similar decrease in blood pressure as a result of the administration of beta-adrenergic receptor and calcium channel blockers does not protect from brain ischemia. 7. In addition, sustained AT(1) receptor inhibition enhances AT(2) receptor expression, associated with increased eNOS activity and NO formation followed by enhanced vasodilatation. Direct AT(1) inhibition and indirect AT(2) receptor stimulation are associated factors normalizing cerebrovascular compliance, reducing cerebrovascular inflammation and decreasing the vulnerability to brain ischemia.8. These results strongly suggest that inhibition of AT(1) receptors should be considered as a preventive therapeutic measure to protect the brain from ischemia, and as a possible novel therapy of inflammatory conditions of the brain.     

Regional cerebral blood flow changes during hypothermia

1. 1. When brain temperature was decreased from 38 to 22 °C using selective hypothermia, tissue blood flow decreased significantly in cerebral cortex, cerebellum, and thalamus, but did not significantly change in hypothalamic or brain stem tissue.

2. 2. A further decrease in brain temperature to 8 °C produced an increase in blood flow in all tissues except cerebral cortex compared to tissue blood flow measured at 22 °C. Compared to normothermic values, blood flow remained significantly decreased at 8 °C in cerebral and cerebellar cortex and was increased in brain stem.

3. 3. After rewarming, tissue blood flow returned to original baseline values in all tissues except cerebral cortex where blood flow was slightly but significantly decreased and brain stem, where blood flow was increased.

4. 4. These results indicate that the cerebrovascular effects of selective brain cooling are regionally specific. These changes appear to be due to both direct and indirect effects of cerebral hypothermia since brain tissue blood flow changes are apparent, compared to control values, after rewarming of the brain.