Dissertations / Theses on the topic 'Brain parenchyma'

2009/2010
In infants and in the Crigler-Najjar syndrome type I patients, severe hyperbilirubinemia due to high levels of unconjugated bilirubin (UCB) may cause Kernicterus, leading to an irreversible and selective brain damage. The Gunn rat is the animal model for the study of these pathologies. It has been suggested that different enzymes of the phase I (cytochrome P-450-dependent mixed function oxygenases 1A1, 1A2, 2A3), phase II (glutathione-S-transferases α2, α3, µ3, µ4, π) and phase III transporters (particularly, Mrp1) seems to be involved in UCB detoxification pathways. However, to date, their in vivo brain expression has been evidenced only at the blood-brain interfaces, while remains largely unexplored in brain parenchyma. Particularly for Mrp1, in vitro evidence reported a role of this transporter in protection of neural primary cultures from dissected cortex, by extruding bilirubin out of the cell. The aim of this study is establish the developmental profile of these genes in brain parenchyma, and assess their alteration in hyperbilirubinemic jj animals. Due to the high regional selectivity of UCB-induced neurotoxicity, cerebellum (Cll), striatum (St), hippocampus (Hip) and cerebral cortex (Cx) were chosen for this study. Our results regard the Mrp1 protein in cerebral cortex of normobilirubinemic (JJ) rats showed that its expression varied during the post-natal age, reaching the highest levels at 9 days after birth. No changes were found between JJ and Jj (having a temporary hyperbilirubinemia in the first week of life) rat for all ages analyzed. Similarly, no differences were detected among JJ/Jj and jj (hyperbilirubinemic) rats at P2, P17 and P60, while a significant increase (p < 0.005) was evidenced in P9 jj rats as compared to age-matched JJ animals. Our Mrp1 mRNA analysis in four regions of P9 animals by Real Time-qPCR revealed the absence of differences among Cx, Cll, St and Hip of P9 normobilirubinemic JJ rats. Moreover, no variations between jj and JJ control animals were detected. Regarding the Mrp1 protein expression in the same four regions by Western blot analysis, our results showed that the levels of this transporter in normobilirubinemic JJ rats were lower in Cx, similar in Cll, St and Hip (p < 0.05 vs Cx). Comparing genotypes, a reduction on Mrp1 in jj animals (compared to Mrp1 amount in the same region of JJ pups) was detected in Cll, St, but reached the statistical significance only in Hip (p < 0.05 vs Hip JJ). The analysis of CYPs gene expression in P9 Gunn rats indicate that CYP1A1, 1A2 and 2A3 mRNA were differently expressed among Cx, Cll, St and Hip of JJ rat. Similarly a region-specific modulation of CYPs expression in jj Gunn rats (compared to JJ) was pointed-out. Surprisingly, UCB seems to generate a plateau effect on CYPs mRNA levels among brain regions of jj rats. In P60 JJ Gunn rats the CYPs expression is higher than in P9 animals, with the following pattern among regions: Cx  CllSt  Hip. A down-regulation (p < 0.05) in St of P60 jj compared to normal animals was observed. Analyzing the GSTs expression in P9 animals, higher variability in the GSTs expression among the four brain areas was evidenced. In hyperbilirubinemic (jj) rats (compared to JJ), statistically relevant down-regulations were detected for GSTα2 (in St;p < 0.05), GSTα3 (in Hip;p < 0.05), µ3 (in Cx;p < 0.01), µ4 (in Cx; p < 0.05) and π (in Cll: p < 0.05); while GSTµ4 was up-regulated in St (p < 0.05). From P9 to P60, in JJ animals: GSTα3 expression increased (13-75-fold depending on the region); while GSTα2 (5-fold), µ3 (p < 0.05), µ4 (2-fold) and π (2-fold) mRNA amounts decreased. In P60 jj Gunn rats, compared to controls (JJ):a relevant up-regulation of GSTα3 was observed in Cll (p < 0.005) and Hip (p < 0.05), while GSTµ3 in jj was down-regulated (p < 0.05). The Mrp1 results obtained in the present in vivo study seems not to be in agreement with the in vitro data reported, to date. Thus, the Mrp1 expression is low in brain parenchyma and bilirubin affect (up-regulation) only marginally the protein amounts in cortex of P9 animals, while in other regions Mrp1 is not modulated, indicating a marginal role in vivo in bilirubin clearance. Similarly, while in liver GSTα2 and α3 act together as ligandin, this seems not happens in brain where the two subunit are expressed at very low levels (P9: α2 77000- α3 1500 fold difference; P60: α2 112000-α3 2200 fold difference with respect to age matched livers). For all genes under analysis, a very complex and variable pattern of expression among brain areas was evidenced. Consequently, no general rules concerning bilirubin-induced modulation could be drawn, as both up and down-regulation were observed. Additionally, in Cx of P9 jj animals, a translational control of Mrp1 might be hypothesized due to a significant increase in Mrp1 protein, without changes in mRNA level. Therefore, the genomic screening made in this work provides the first general overview on the mRNA developmental profiles of several CYPs and GSTs genes in brain parenchyma (specifically Cx, Cll, St and Hip) of normal rats, and of animals suffering from hyperbilirubinemia, underlying the necessity to find functional evidence to finally understand the role of these enzymes associated with the kernicterus and Crigler-Najjar type I syndrome pathologies.
XXIII Ciclo
1978

Introduction: Accumulation of the β-amyloid (Aβ) protein in cerebral blood vessels is a hallmark of Alzheimer’s disease. Identifying the factors that contribute to this accumulation is critical for both prevention and treatment of the disease. Previous studies have shown that Aβ from the parenchyma is removed along the basement membranes of capillaries and arteries towards the surface of the brain. Aβ is present in the cerebrospinal fluid (CSF) of the brain and there have been suggestions that Aβ is entering the parenchyma and eliminated back into the CSF. The precise anatomical routes by which CSF moves into and out of the brain parenchyma have not been identified. The aim of this study was to determine the entry route and the distribution of solutes and particulate matter in the brain at two time points following its injection into the CSF and investigate age-related changes in CSF influx and distribution. Materials and method: Evans blue dye (EBD), anti-sense oligonucleotides (ASO), fluorescently-tagged Aβ40 and nanoparticles were injected into cisterna magna of adult male C57BL/6J mice and mice were sacrificed at 5 or 30 minutes after injection (n = 3 /group). Representative sections across the brain were immunostained for the detection of basement membranes and smooth muscle actin to differentiate arteries from veins by confocal microscopy. Nanoparticles were detected using transmission electron microscopy. Results and discussion: Within 5 minutes after injection, EBD, ASO, nanoparticles and Aβ entered the brain along the pial – glial vascular basement membranes. In 6 – 10 week old mice, soluble Aβ colocalised with the astrocytic basement membrane marker (α-2 laminin) and collagen IV of leptomeningeal and cortical arteries. After 30 minutes, Aβ was present in the tunica media of leptomeningeal and cortical arteries. The depth of Aβ along the artery wall in the parenchyma was significantly higher at 30 min compared to 5 min post-injection. Regional differences between the cortical, subcortical and posterior brain regions were also detected. In 24 – 30 month old mice, Aβ also entered along the pial – glial basement membrane of arteries after 5 min. However, after 30 min, Aβ was also present in 1) the basement membrane of capillaries, veins, 2) the tunica media of arteries, 3) diffusely in the parenchyma where it was taken up by astrocytes, neurons and macrophages. Conclusions: These results indicate that the entry route of CSF is along the pial – glial basement membrane of arteries at 5 min post-injection. Solutes in the CSF appear to enter the intramural periarterial drainage pathway after 30 min for clearance back into the CSF. The normal flow of CSF along the basement membrane of arteries in old mice is impaired. These data may contribute to the understanding of Aβ deposition in walls of arteries as cerebral amyloid angiopathy and provide an insight into the intrathecal delivery route for treatment of neurodegenerative diseases including Alzheimer’s disease.

In certain diseases, like Multiple Sklerosis and Alzheimer's disease, the progression of the disease can be measured by whole brain atrophy. A difficulty with this is that all people have very different scull sizes, thus also very different brain sizes. This makes it almost impossible to establish "normal values" for brain size. The spread is very large and the method is not practical to use for individual patients. A method with less spread in healthy persons is to use the ratio of the Brain Parenchymal Fraction (BPF). The use of Brain Parenchymal Fraction has increased steadily since it was first introduced in 1999.  BPF = BPV/ICV This study was performed to increase the knowledge of what is normal and to evaluate the robustness of the BPF as a measurement for brain atrophy. Among other things, the change in the BPF when calculated from incomplete volumes (parts of the scull missing in the set of MR images) was evaluated.  The results show that when parts are missing from the top (superior) of the scull the resulting BPF is strictly higher than the correct PBF and when parts are missing from the lower (inferior) part of teh scull the resulting BPF is stritly lower than teh correct value.  Two different methods where tried to compensate for missing parts. The first method was to find a variable factor to compensate with, the size of this factor was depending on how much of the scull that was missing. The second method was to interpolate the ICV and BPV curves and from the new interpolated curves, calculate a new BPF. The method of compensating incomplete volumes using a factor calculated as a function of the intercranial volume of the first/last available slice turned out to be the better.

Background Multiple sclerosis (MS) is an autoimmune disease characterised by inflammatory damage to the central nervous system (CNS). Accumulated CNS injury can be quantified as brain atrophy, definable as a reduction in brain parenchymal fraction (BPF). BPF correlate with disability in MS and is used routinely as an endpoint in clinical trials. In 2009/2010, a new MS clinical care program, that includes follow-up of BPF, was introduced at Umeå University Hospital (NUS). Levels of neurofilament light polypetide (NFL) and glial fibrillary acidic protein (GFAP) in cerebrospinal fluid (CSF) are markers of axonal and astrocytic injury, respectively, and also potential surrogate biomarkers for BPF decline. The goals of this thesis were to establish age-adjusted values of BPF in healthy individuals and to relate these to the BPF values from individuals with MS as well as to the levels of NFL and GFAP in CSF. Another goal was to investigate if expanded disability status scale (EDSS)-worsening could be predicted in a clinical MS cohort and if BPF measurements could contribute to such predictions. Methods A group of 111 healthy individuals volunteered to participate in the studies. A total of 106 of these underwent MRI with BPF measurements, 53 underwent lumbar puncture (LP) with measurement of NFL and GFAP and 48 underwent both MRI and LP. Three different automatic and one manual method were utilised to determine BPF. A literature search on BPF in healthy individuals was performed for the purpose of a systematic review. For studying disability progression in MS, all individuals with MS followed at NUS and included in the Swedish MS registry were included if they had matched data on BPF, EDSS and lesion load as part of clinical follow-up (n=278). Results BPF as well as NFL and GFAP levels in CSF were all associated with age. NFL was associated with BPF and GFAP, but only the association with GFAP was retained when adjusting for age. Significant differences were found between different methods for BPF determination. In the MS population, BPF was associated with EDSS. Only progressive disease course could predict EDSS worsening. Conclusion The data on BPF and levels of NFL and GFAP in CSF of healthy individuals can aid in the interpretation of these variables in the setting of MS. Knowledge on differences in BPF data from different methods for BPF determination can be useful in comparing data across studies, but also highlights the need for a commonly accepted gold standard. The correlation between GFAP and NFL levels in CSF may indicate an association between glial and axonal turnover that is independent of the aging effect on the brain. However, the low number of volunteers for LP precluded clear conclusions. An association between BPF and EDSS was seen in the MS group. The ability to predict EDSS worsening in the clinical MS cohort was limited.

In certain neuro-degenerative diseases likemultiple sclerosis (MS), the rate of brain atrophy can be measured by monitoring the brain parenchymal fraction (BPF) in such patients. The BPF is defined as the ratio of brain parenchymal volume (BPV, defined as the total volume of gray matter tissue, white matter tissue and other unidentified tissue) and intracranial volume (ICV, the total volume of the skull). It can be represented by the formula in equation 1: A complication with this measure is that the BPF is affected by the presence of edema in the brain, which leads to swelling and hence may obscure the true rate of brain atrophy. This leads to uncertainty when establishing “normal values” of BPF when analyzing different magnetic resonance imaging (MRI) scans of the same patient. Another problem is that different MRI scans of the same patient cannot be compared directly, due to the fact that the head of the patient will be in a different position for every scan. The SyMRI software used in this master thesis has the functionality to perform brain tissue characterization and measurement of brain volume, given a number of MR images of a patient. Using tissue properties such as longitudinal relaxation time (T1), transverse relaxation time (T2) and proton density (PD), each voxel in a volume can be classified to belong to a certain tissue type. From these measurements, the intracranial volume, brain volume, white matter, gray matter and cerebrospinal fluid volumes can easily be estimated. In this master thesis, the BPF of several patients were analyzed based on quantitative MRI (qMRI) images, in order to identify the change of BPF due to the presence of edema over time. Volumes obtained from the same patients at different time points were aligned (registered), such that the BPF can be easily compared between years. A correlation analysis between the BPF and R1, R2 and PD was performed (R1 is the longitudinal relaxation rate defined as 1/T1 relaxation time and R2 Is transverse relaxation rate defined as 1/T2 relaxation time) to investigate if any of these variables can explain the change in BPF. The results show that due to image registration, and removing some of the slices from the top and bottom of the head, the BPF of the patients was corrected to a certain extent. The change in the mean BPF of each patient over four years was less than 1% post registration and slice removal. However, the decrease in standard deviation was between 6.9% to 52% after registration and removing of slices. The BPF of the follow-up years also came closer to the initial BPF value measured in the first year. The statistical analysis of the BPF and R1, R2 and PD, showed a very low correlation (0.1) between BPF and PD, and intermediate correlations between BPF and R1, R2 (0.385 and -0.51, respectively). Future work will focus on understanding how these results relate to edema.

Despite previous beliefs, the generation of new neurons and new glia in the central nervous system (CNS) continues throughout life. Adult neurogenesis occurs in both classical neurogenic niches (e.g., the subventricular zone, SVZ, of the lateral ventricles) as well as in the entire brain's parenchyma, which is full of quiescent neural progenitors that are activated after injury. In particular, NG2-positive polydendrocytes, that usually differentiate to mature oligodendrocytes participating to re-myelination after injury, retain some multipotency and, under some conditions, can also generate neurons and astrocytes (Nishiyama et al., 2009). Among various neurotransmitters and growth factors, extracellular nucleotides (ATP, UTP, their break-down products and sugar nucleotides), which are released at high amounts at the sites of CNS damage (Ulrich et al., 2012) are key actors in regulating reparative responses via purinergic P2 receptors (Abbracchio et al., 2009). Previous studies from our laboratory have identified the purinergic receptor GPR17 as a new marker of early stages of NG2 cell differentiation, showing that GPR17 activation accelerates NG2 cells’ oligodendrocyte fate (Fumagalli et al., 2011; Ceruti et al., 2011; Boda et al., 2011). Interestingly, GPR17 is also one of the key genes expressed by human adult neural stem cells (Maisel et al., 2007), suggesting a possible role in cell fate determination. Based on these premises, the aim of my PhD project was to investigate the role of purinergic signaling in regulating stem cell proprieties of adult brain subventricular zone and of NG2+ parenchymal progenitors. In the first part of my PhD thesis, by using a conditional GLAST::CreERT2 Rosa YFP mouse model and an in vitro neurosphere assay, we have demonstrated that the P2Y receptor agonist ADPβS promotes the proliferation of SVZ neural progenitors and sustains their progression towards the generation of neuroblasts, either directly or through the activation of parenchymal astrocytes. In the second part of my PhD, OPCs have been then cultured accordingly to two published protocols (Kondo & Raff, 2000; Liu et al., 2007) both able to unveil their stem cell properties. In both protocols, we observed an increase in the percentage of cells expressing the neuronal marker β-tubulinIII (βtubIII) upon treatment with the non-selective GPR17 antagonist Cangrelor and VPA with respect to control. Interestingly GPR17, which is normally expressed only by NG2+ cells, was surprisingly detected in a subset of βtubIII+ cells already under control differentiative condition, suggesting its potential involvement in neurogenesis. The appearance of this cell population was further incremented by the exposure to VPA and Cangrelor. Taken together, our results suggest that antagonizing GPR17 functions can address the fate of NG2 cells towards the generation of new neurons, as observed with epigenetic modulators like VPA. In conclusion, our results strengthen the evidence that the purinergic system crucially regulates neuronal progenitors, either in a classical neurogenic niche or in the brain parenchyma. The pharmacological modulation of the purinergic system could therefore represent a promising and innovative approach to exploit the intrinsic ability of the adult brain to regenerate in acute and chronic neurodegenerative disorders.

Tese de mestrado, Ciências Biofarmacêuticas, Universidade de Lisboa, Faculdade de Farmácia, 2016
Despite the restricted permeability of the blood-brain barrier (BBB), the brain is a privileged organ regarding the appearance of metastases, particularly from breast cancer. Patients with brain metastases from breast cancer have a severe prognosis, rendering this issue a serious oncologic problem that deserves further attention. Therefore, additional studies are required to establish when breast cancer cells cross the brain endothelium and what are the routes used for the transendothelial migration, to understand what is their precise phenotype along the processes of transmigration and establishment of brain metastases, to determine the alterations occurring in brain endothelium, to study how endothelial cells communicate with malignant ones to promote the attraction of malignant cells into the brain vasculature and tumour-associated vascular development. Based on this, we aimed at establishing the temporal profile of breast cancer metastasization to the brain and characterize the metastasizing cells phenotype, as well as, to investigate the vascular events and BBB properties along the process of metastasization to this target organ. In addition, we aimed to assess signalling mechanisms involved in attraction of carcinoma cells into the brain and proliferation in the nervous tissue. To establish the temporal evolution of the players involved in such processes, we used cerebella, cranial hippocampi, and striata of female mice inoculated with 4T1 breast cancer cells sacrificed at 5 hours, 3 days, 7 days or 10 days, and of female mice injected with vehicle (control) sacrificed at 5 hours. Our results showed the presence of brain metastasis of breast cancer at 7-days after inoculation, which increased thereafter. The malignant cells crossed the BBB as mesenchymal cells and, once inside the brain, these cells underwent a complete or partial mesenchymal-epithelial transition to acquire the epithelial characteristics that allow the establishment of new tumours. In addition, the process of brain metastasization of BC contributed to the downregulation of the tight junction protein claudin-5 of brain microvascular endothelial cells, as well as to the entrance of the blood-borne component thrombin in brain parenchyma. On the other hand, hypervascularization in cranial hippocampus appeared to be associated to the process of brain colonization by breast cancer cells. Regarding the role of platelet-derived growth factor B signalling along the process of brain metastasization, we found that this growth factor was expressed by tumour cells and its expression increased during the formation of brain metastasis. Interestingly, a continuous entrance of cysteine-X amino acid-cysteine receptor 4 (CXCR4)-positive cells into the brain parenchyma appeared to occur along the process of brain metastasization of breast cancer. In sum, this study contributes to clarify the time-course and interdependence of the signalling events, BBB breach and phenotypic transition of malignant cells along endothelial transposition and brain metastases establishment by breast cancer cells. Moreover, the demonstration of early cellular and molecular events points to novel targets for modulation in order to prevent metastasis formation and development.
Apesar da barreira hematoencefálica (BHE) ter uma permeabilidade restrita, o cérebro é um órgão preferencialmente afetado pelo aparecimento de metástases, particularmente de cancro mama. Pacientes com metástases cerebrais provenientes do cancro da mama têm um prognóstico severo, tornando a metastização num sério problema oncológico que merece toda a atenção. Por este motivo, novos estudos são necessários para estabelecer quando é que as células cancerígenas da mama atravessam o endotélio cerebral e quais são as vias que utilizam para migrarem através do endotélio, para se perceber qual o fenótipo que têm ao longo dos processos de migração para dentro do encéfalo e durante o estabelecimento de metástases, para determinar as alterações que ocorrem no endotélio cerebral, para estudar como as células endoteliais comunicam com as células malignas para promover a atracão das células cancerígenas da mama para a vasculatura do encéfalo e o desenvolvimento vascular associado ao tumor. Com base nisto, tivemos com objetivos estabelecer o perfil temporal da metastização do cancro da mama para o encéfalo e caracterizar o fenótipo destas mesmas células, assim como, estudar as alterações vasculares e as propriedades da BHE ao longo do processo de metastização para este órgão secundário. Para além disso, também pretendíamos avaliar os mecanismos de sinalização envolvidos na atracão das células tumorais para o encéfalo e na proliferação no tecido nervoso. Para estabelecer a evolução temporal dos intervenientes envolvidos em tais processos, utilizámos cerebelos, hipocampos craniais e estriados de ratinhos fêmea inoculados com células cancerígenas da mama 4T1 sacrificados às 5 horas, 3 dias, 7 dias, ou 10 dias, e de ratinhos fêmea injetados com veículo (controlo) sacrificados às 5 horas. Os nossos resultados mostraram a presença de metástases cerebrais do cancro mama 7 dias após a inoculação, aumentando ao longo do tempo. As células malignas atravessaram a BHE como células mesenquimais e, uma vez dentro do encéfalo, estas células sofreram uma transição completa ou parcial de fenótipo mesenquimal para epitelial para adquirirem as características epiteliais necessárias para o estabelecimento de novos tumores no encéfalo. Além disso, o processo de metastização cerebral do cancro da mama contribuiu para a diminuição da expressão da proteína das junções de oclusão claudina-5 nas células endoteliais microvasculares cerebrais, assim como para a entrada do componente sanguíneo trombina no parênquima cerebral. Por outro lado, o aumento de vascularização no hipocampo cranial aparentou estar associado ao processo de colonização do encéfalo pelas células cancerígenas da mama. Relativamente ao papel da sinalização do fator de crescimento B derivado de plaquetas ao longo do processo de metastização cerebral, descobrimos que as células tumorais expressavam este fator de crescimento e que a sua expressão aumentou durante a formação de metástases no encéfalo. Curiosamente, a entrada contínua de células que expressam o recetor CXCR4 para dentro do parênquima cerebral aparentou ocorrer ao longo do processo de metastização cerebral do cancro da mama. Deste modo, este estudo contribui para clarificar o curso temporal e a interdependência de vias de sinalização, a quebra da BHE e a transição fenotípica das células malignas ao longo na transposição do endotélio e estabelecimento de metástases cerebrais pelas células cancerígenas da mama. Além disso, a demonstração dos eventos celulares e moleculares iniciais aponta para novos alvos para modulação de modo a prevenir a formação e desenvolvimento de metástases.
This study was supported by Fundação para a Ciência e Tecnologia (FCT – UID/DTP/04138/2013), Portugal, and by National Research, Development and Innovation/Hungarian Scientific Research Fund (NKFIH/OTKA – K-100807 and K-116158), Hungary.

碩士
國立勤益科技大學
電子工程系
98
Making use of magnetic field theory, magnetic resonance imaging (MRI) which employs pulses radio waves and intense magnetic fields to vibrate hydrogen atoms in the body is a method of obtaining computer images. After long-term clinical trials, MRI has been proved to use in humans harmlessly because of its advantages of non-invasive and non-radiation. In addition, MRI provides cross-sectional images of the objective and scans with different frequency. In light of the procedure, it will generate the difference in strength characteristics, combining powerful imaging sequence changes to provide abundant organization information, which is the key to use computer algorithms to reduce the burden on physician interpretation. How to use computer to decrease processing time is the point, comparing to burden of the physicians diagnosis. In this study, the automatic vector seed region growing is applied in the brain MRI classification. First, vector seed is selected automatically. Second, base on seed regional growth, brain is segmented in detailed. Finally, using K-mean to classify the brain segmentation images, according to gray matter, white matter, and cerebral spinal fluid. When physicians face many images, automatic analysis of medical images become the most challenging task. Nowadays automatic vector seed region growing which is proposed can be the reference for many researchers because it can decrease processing time particularly. This approach makes physicians’ diagnosis acute and effectiveness. Compare to K-means and FMRIB’s Automated Segmentation Tool, FAST, the classified statistics of AVSRG is above all else. To prove the automatic classification results Vector seeds of regions growing, using the receiver operating characteristic analysis to evaluate classification performance, and prove its superiority.

碩士
國立陽明大學
生物醫學影像暨放射科學系
102
Background: Brain perfusion parameters are altered by the presence of stenosis or occlusion of internal carotid artery. A lesion can’t be distinguished from normal tissue by using a single perfusion parameter perfectly because the parameter values varied with the components of different brain tissues. The distinct distributions of lesion and normal brain parenchyma in a scatter plot with two parameters can be separated by support vector machine (SVM) with an optimal dividing line. According to the algorithm, the mask of normal brain parenchyma can be identified. Material and method: We retrospectively analyzed 7 cases with severe unilateral internal carotid artery stenosis. Six combinations of perfusion parameters were choosing from all scatter plots for classification by SVM. Sifting the best combination for SVM to identify normal brain parenchymal mask, and then the mean values of perfusion parameters of the mask calibrated with standard values from literature. The scaling factors were applied to whole brain to calculate the absolute perfusion parameters. Two different approaches for identification of normal brain masks, the contralateral cerebral hemisphere and the median of time to peak (TTP), were compared with SVM to evaluate the accuracy of normal brain masks. Result: Tmax-TTP scatter plot provided the best result with classification by SVM. The percentage of difference between absolute values of perfusion parameters and standard values demonstrated the superiority of SVM. Conclusion: The accuracy of normal brain mask identified by SVM can improve the absolute values of brain perfusion parameters.

Yes
To determine whether or not jugular venous reflux (JVR) is associated with structural brain parenchyma changes in individuals with mild cognitive impairment (MCI) and Alzheimer's disease (AD). 16 AD patients (mean (SD): 81.9 (5.8) years), 33 MCI patients (mean (SD): 81.4 (6.1) years) and 18 healthy elderly controls (mean (SD): 81.5 (3.4) years) underwent duplex ultrasonography and magnetic resonance imaging scans to quantify structural brain parenchyma changes. Normalized whole brain (WB), gray matter (GM) and white matter (WM) volumes were collected, together with CSF volume. JVR was strongly associated with increased normalized WB (p = 0.014) and GM (p = 0.002) volumes across all three subject groups. There was a trend towards increased WB and GM volumes, which was accompanied by decreased CSF volume, in the JVR-positive subjects in both the MCI and AD groups. When the MCI and AD subjects were aggregated together significant increases were observed in both normalized WB (p = 0.009) and GM (p = 0.003) volumes for the JVR-positive group. No corresponding increases were observed for the JVR-positive subjects in the control group. Through receiver operating characteristic analysis of the brain volumetric data it was possible to discriminate between the JVR-positive and negative AD subjects with reasonable accuracy (sensitivity = 71.4%; specificity = 88.9%; p = 0.007). JVR is associated with intracranial structural changes in MCI and AD patients, which result in increased WB and GM volumes. The neuropathology of this unexpected and counterintuitive finding requires further investigation, but may suggest that JVR retrogradely transmits venous hypertension into the brain and leads to brain tissues swelling due to vasogenic edema.

The direct endothelial contact with adjacent astrocytic end-feet is believed to establish blood-brain barrier (BBB) typical characteristics in endothelial cells of the central nervous system (CNS). However, this contact is only present in capillary vessels of the brain parenchyma and absent in larger veins, arteries and vessels within the meninges. To investigate a potential impact of direct endothelial interactions with adjacent astrocytic end-feet on the molecular tight junction (TJ) composition and ultrastructure, we performed a systematic analysis of endothelial cell contacts within the vascular tree of parenchymal and leptomeningeal vessels. Immunofluorescence labeling for claudin-3, claudin-5, zonula occludens-1 and occludin was used to compare the molecular composition, without showing significant differences in their distribution along the vascular tree of parenchymal and leptomeningeal vessels. Furthermore, electron microscopy in combination with quantitative analyses was performed to investigate the endothelial ultrastructure revealing significant differences within the length of endothelial overlaps between the different vessel types. Here, parenchymal arteries exhibit noticeably longer cell contacts compared to capillaries, which could not be observed in leptomeningeal vessels. It was also observed that arterial vessels regularly contain a higher density of endothelial vesicles throughout the parenchyma and meninges as a sign for transendothelial traffic. Hence, endothelial expression of blood-brain barrier typical TJs is not limited to capillary vessels with an intimate contact to surrounding astrocytes, but is also observed in arteries and veins of the brain parenchyma as well as the meninges, the latter of which are lacking a direct astrocyte-endothelial interaction. These vessel-specific characteristics can now be used to address and compare alterations of the BBB in different settings of CNS pathologies.:Table of Content 1. INTRODUCTION 4 1.1 THE BLOOD-BRAIN BARRIER 4 1.2 HISTORY 5 1.3 STRUCTURE AND COMPOSITION 6 1.4 THE ROLE OF THE MICROENVIRONMENT 8 1.4.1 ASTROCYTES 8 1.4.2 PERICYTES 9 1.5 BLOOD BRAIN BARRIER FUNCTION 10 1.5.1 PHYSIOLOGIC CONDITIONS 10 1. 5.2 PATHOLOGIC CONDITIONS 11 2. OPEN QUESTIONS AND SCIENTIFIC APPROACH 12 3. PUBLICATIONS 13 3.1 DIFFERENT SEGMENTS OF THE CEREBRAL VASCULATURE REVEAL SPECIFIC ENDOTHELIAL SPECIFICATIONS, WHILE TIGHT JUNCTION PROTEINS APPEAR EQUALLY DISTRIBUTED 13 3.2 THE BLOOD-BRAIN BARRIER 28 4. SUMMARY 40 5. REFERENCES 43 6. PROOF OF SIGNIFICANT CONTRIBUTION 48 7. DECLARATION OF ACADEMIC HONESTY 49 8. ACKNOWLEDGMENT 50 9. CURRICULUM VITAE 51