A Novel Quantification Method for Determining Previously Undetected Silent Infarcts on MR-perfusion in Patients Following Carotid Endarterectomy

The purpose of this paper is to evaluate the post-operative Magnetic Resonance Perfusion (MRP) scans of patients undergoing carotid endarterectomy (CEA), using a novel image-analysis algorithm, to determine if post-operative neurocognitive decline is associated with cerebral blood flow changes. CEA procedure reduces the risk of stroke in appropriately selected patients with significant carotid artery stenosis. However, 25% of patients experience subtle cognitive deficits after CEA compared to a control group. It was hypothesized that abnormalities in cerebral blood flow (CBF) are responsible for these cognitive deficits. A novel algorithm for analyzing MRperfusion (MRP) scans to identify and quantify the amount of CBF asymmetry in each hemisphere was developed and to quantify the degree of relative difference between three corresponding vascular regions in the ipsilateral and contralateral hemispheres, the Relative Difference Map (RDM). Patients undergoing CEA and spine surgery (controls) were examined preoperatively, and one day postoperatively with a battery of neuropsychometric (NPM) tests, and labeled “injured” patients with significant cognitive deficits, and “normal” if they demonstrated no decline in neurocognitive function. There are apparently significant RDM differences with MRP scans between the two hemispheres in patients with cognitive deficits which can be used to guide expert reviews of the imagery. The proposed methodology aids in the analysis of MRP parameters in patients with cognitive impairment

Enhanced Techniques for Asymmetry Quantification in Brain Imagery

We present an automated generic methodology for symmetry identification and asymmetry quantification, novel method of identifying and delineation of brain pathology by analyzing the opposing sides of the brain utilizing of inherent leftright symmetry in the brain. After symmetry axis has been detected, we apply non-parametric statistical tests operating on the pairs of samples to identify initial seeds points which is defined defined as the pixels where the most statistically significant difference appears. Local region growing is performed on the difference map, from where the seeds are aggregating until it captures all 8-way connected high signals from the difference map. We illustrate the capability of our method with examples ranging from tumors in patient MR data to animal stroke data. The validation results on Rat stroke data have shown that this approach has promise to achieve high precision and full automation in segmenting lesions in reflectional symmetrical objects

Added value of acute multimodal CT-based imaging (MCTI) : a comprehensive analysis

Introduction: MCTI is used to assess acute ischemic stroke (AIS) patients.We postulated that use of MCTI improves patient outcome regardingindependence and mortality.Methods: From the ASTRAL registry, all patients with an AIS and a non-contrast-CT (NCCT), angio-CT (CTA) or perfusion-CT (CTP) within24 h from onset were included. Demographic, clinical, biological, radio-logical, and follow-up caracteristics were collected. Significant predictorsof MCTI use were fitted in a multivariate analysis. Patients undergoingCTA or CTA&CTP were compared with NCCT patients with regards tofavourable outcome (mRS ≤ 2) at 3 months, 12 months mortality, strokemechanism, short-term renal function, use of ancillary diagnostic tests,duration of hospitalization and 12 months stroke recurrence

Stroke, atrial fibrillation, and the management of oral anticoagulation

Atrial fibrillation (AF) is the most common arrhythmia, becomes more prevalent with increasing age and is linked to neurological complications, including most notably ischemic stroke, but also cognitive dysfunction. The recent introduction of direct oral anticoagulants (DOAC) significantly advanced the management of patients with AF. Large-scale randomized controlled trials showed that – for most patients – DOAC are at least as effective as vitamin K antagonists (VKA) in preventing ischemic stroke but have the advantage of a lower risk for intracranial hemorrhage (ICH). However, clinically important patient populations were underrepresented or excluded in these trials, and more refined aspects which are important in clinical practice, including concomitant stroke etiologies or brain pathologies, medication adherence, neuroimaging characteristics or biomarker signatures were not addressed. Therefore, several research gaps and challenges remained for neurologists treating patients with AF, of which we selected 4 aspects to focus on in the following topics that comprise this PhD thesis. The first topic focused on 3 high-risk subgroups of patients with AF treated with oral anticoagulants who were underrepresented or excluded from the large randomized trials. These were (i) stroke patients aged 85 years and older (“the oldest-old”), (ii) severely affected stroke patients dependent on the daily help of others, and (iii) stroke patients with concomitant cerebral small vessel disease. In the first main project of this PhD thesis we examined the performance of DOAC versus VKA in the oldest-old patients with recent stroke and AF in a large pooled analysis across 7 cohort studies. Facing the paucity of randomized evidence, many physicians have been reluctant to use DOAC in these patients. With this project, we provided new evidence that the benefits of DOAC over VKA are preserved in the oldest old with recent stroke, without any signal of a safety concern regarding risk of ICH. In an additional project from our local cohort, we showed that the favourable profile of DOAC over VKA was preserved also among patients with AF and recent stroke who were dependent on the daily help of others, a patient subgroup for which no data existed previously. Finally, we showed in the same cohort that concomitant cerebral small vessel disease in anticoagulated patients with AF and recent stroke was associated with an unfavorable clinical course, but the risk for ischemic stroke remained higher than the risk for ICH, even in the presence of small vessel disease. The latter two projects fall within the scope of this PhD thesis, but do not constitute its main body. Although both manuscripts were published in peer-reviewed journals, for the purpose of this thesis their presentation is restricted to abstracts. Our findings in the first topic of this PhD thesis advance the evidence for the use of anticoagulants in high-risk patient subgroups with AF and recent stroke. The second topic examined ischemic stroke occurring despite anticoagulant therapy in AF patients. With the increasing use of oral anticoagulants, this scenario represents a growing challenge in everyday clinical practice, indicating the need to elucidate the underlying causes and – based on these – the optimal subsequent management strategies. We addressed this issue in the second main project of this PhD thesis in a large retrospective analysis pooling data of prospectively collected patients from 11 stroke centers. We found that the causes of stroke despite anticoagulation in AF patients were heterogeneous, but form three main clusters, all of which were comparably important. These included (i) competing stroke mechanisms other than AF-related cardioembolism, and (ii) insufficient anticoagulation due to prescription errors and nonadherence, suggesting that individualized treatment approaches to address these causes are necessary. Importantly, the third and most common cause was AF-related cardioembolism despite sufficient anticoagulation, indicating the need to develop novel preventive strategies beyond the currently available anticoagulants. Furthermore, in this project we were able to demonstrate that AF patients with stroke despite anticoagulation represent a high-risk patient population, with higher than expected rates of stroke recurrence and other unfavorable outcomes. Finally, we showed also in this population that subsequent treatment with DOAC was associated with better outcomes than VKA treatment. Interestingly, neither any specific switch between DOAC nor antiplatelets as add-on treatment to anticoagulation seemed to confer any benefit, although both approaches are often employed in clinical practice. This study advanced the evidence for the preferential use of DOAC over VKA in AF patients with stroke despite anticoagulation, for whom no data existed so far, while demonstrating the need for more individualized and novel treatment approaches in these high-risk patients. The third topic of this PhD thesis was the adherence of stroke patients to DOAC. Unlike VKA, DOAC require no coagulation monitoring and have short half-lives, which has raised concerns about nonadherence in AF patients treated with DOAC. This is particularly pertinent to patients with stroke, as shown in the second topic of this PhD thesis. In order to examine the medication-taking behaviour and the effect of an adherence-enhancing intervention in patients with recent stroke, we designed, initiated, and undertook the MAAESTRO study. MAAESTRO has been a joint venture with the Pharmaceutical Care Research Group of the University of Basel and has used electronic monitoring as the main method to assess adherence, data on which have been scarce so far. MAAESTRO comprises an initial observational phase and a subsequent randomized controlled interventional phase. MAAESTRO successfully concluded recruitment in July 2021 with reaching the predefined goal of n=130 participants. The observational study phase has now been completed, but as follow-up in the interventional phase is ongoing, the main study results are not part of this PhD thesis. Still, we present the published study protocol and the first results from the observational phase on the patterns of DOAC-taking behaviour, as well as an exploratory analysis on how adherence was impacted by the COVID-19 lockdown as abstracts, as these publications fall within the scope of this PhD thesis, but do not formally constitute its main body. The fourth and final topic of this PhD thesis focused on cognitive dysfunction as a neurological complication of AF. While stroke is a well-known consequence of AF, there is increasing evidence that AF is also linked to cognitive dysfunction independent of ischemic stroke, but the mechanisms underlying this association are unclear. To preserve cognitive function in the growing population of elderly AF patients, a better understanding of these mechanisms is needed. Using data from the multicenter Swiss-AF Cohort Study, in the third main project of this PhD thesis we investigated serum neurofilament light chain (sNfL), a novel blood-based biomarker of neuronal damage, as a tool to explore the mechanisms through which neurological disease occurs in AF. In a cross-sectional analysis, we showed that sNfL is inversely associated with brain volume and cognitive function, thereby demonstrating that it represents a relevant biomarker of brain health in AF patients. Furthermore, we showed that neuronal loss measured by sNfL is associated with age, diabetes mellitus, heart failure, blood pressure and vascular brain lesions, observations which provide mechanistic insights into the occurrence of neurological disease in AF. Finally, in the fourth main project of this PhD thesis, we additionally investigated in this elderly cardiovascular cohort how renal function and body mass index contribute to sNfL levels in order to gain insights into the homeostasis (i.e., clearance and distribution) of this biomarker in the blood compartment. A better understanding of this is necessary towards further establishing this neurological biomarker in cardiovascular and dementia research. We showed that both renal function and body mass index were strongly, inversely associated with sNfL, but only renal function explained a relevant proportion of its variance. With this project we provided evidence for the importance of accounting for renal function in future sNfL-based investigations in elderly cardiovascular populations, in whom chronic kidney disease is highly prevalent

Assessment of abdominal aortic aneurysm biology using magnetic resonance imaging and positron emission tomography-computed tomography.

Background Although abdominal aortic aneurysm (AAA) growth is non-linear, serial measurements of aneurysm diameter are the mainstay of aneurysm surveillance and contribute to decisions on timing of intervention. Aneurysm biology plays a key part in disease evolution but is not currently routinely assessed in clinical practice. Magnetic Resonance Imaging (MRI) and Positron Emission Tomography-Computed Tomography (PET-CT) provide insight into disease processes on a cellular or molecular level, and represent exciting new imaging biomarkers of disease activity. Macrophage-mediated inflammation may be assessed using ultrasmall superparamagnetic particles of iron oxide (USPIO) MRI and the PET radiotracer 18FSodium Fluoride (18F-NaF) identifies microcalcification which is a response to underlying necrotic inflammation. The central aim of this thesis was to investigate these imaging modalities in patients with AAA. Methods and Results USPIO MRI: MULTI-CENTRE STUDY In a prospective multi-centre observational cohort study, 342 patients (85.4% male, mean age 73.1±7.2 years, mean AAA diameter 49.6±7.7mm) with asymptomatic AAA ≥4 cm anteroposterior diameter underwent MRI before and 24-36 hours after intravenous administration of USPIO. Colour maps (depicting the change in T2* caused by USPIO) were used to classify aneurysms on the basis of the presence of USPIO uptake in the aneurysm wall, representing mural inflammation. Intra- and inter-observer agreement were found to be very good, with proportional agreement of 0.91 (kappa 0.82) and 0.83 (kappa 0.66), respectively. At 1 year, there was 29.3% discordant classification of aneurysms on repeated USPIO MRI and at 2 years, discordance was 65%, suggesting that inflammation evolves over time. In the observational study, after a mean of 1005±280 days of follow up, there were 126 (36.8%) aneurysm repairs and 17 (5.0%) ruptures. Participants with USPIO enhancement (42.7%) had increased aneurysm expansion rates (3·1±2·5 versus 2·5±2·4 mm/year; difference 0·6 [95% confidence intervals (CI), 0·02 to 1·2] mm/year, p=0·0424) and had higher rates of aneurysm rupture or repair (69/146=47·3% versus 68/191=35·6%; difference 11·7%, 95% CI 1·1 to 22·2%, p=0·0308). USPIO MRI was therefore shown to predict AAA expansion and the composite of rupture or repair, however this was not independent of aneurysm diameter (c-statistic, 0·7924 to 0·7926; unconditional net reclassification -13·5%, 95% confidence intervals -36·4% to 9·3%). 18F-NaF PET-CT: SINGLE-CENTRE STUDY A sub-group of 76 patients also underwent 18F-NaF PET-CT, which was evaluated using the maximum tissue-to-background ratio (TBRmax) in the most diseased segment (MDS), a technique that showed very good intra- (ICC 0.70-0.89) and inter-observer (ICC 0.637-0.856) agreement. Aneurysm tracer uptake was compared firstly in a case-control study, with 20 patients matched to 20 control patients for age, sex and smoking status. 18F-NaF uptake was higher in aneurysm when compared to control aorta (log2TBRmax 1.712±0.560 vs. 1.314±0.489; difference 0.398 (95% CI 0.057, 0.739), p=0.023), or to non-aneurysmal aorta in patients with AAA (log2TBRmax 1.647±0.537 vs. 1.332±0.497; difference 0.314 (95% CI 0.0685, 0.560), p=0.004). An ex vivo study was performed on aneurysm and control tissue, which demonstrated that 18F-NaF uptake on microPET-CT was higher in the aneurysm hotspots and higher in aneurysm tissue compared to control tissue. Histological analysis suggested that 18F-NaF was highest in areas of focal calcification and necrosis. In an observational cohort study, aneurysms were stratified by tertiles of TBRmax in the MDS and followed up for 510±196 days, with 6 monthly serial ultrasound measurements of diameter. Those in the highest tertile of tracer uptake expanded more than 2.5 times more rapidly than those in the lowest tertile (3.10 [3.58] mm/year vs. 1.24 [2.41] mm/year, p=0.008) and were also more likely to experience repair or rupture (15.3% vs. 5.6%, log-rank p=0.043). In multivariable analyses, 18F-NaF uptake on PET-CT emerged as an independent predictor of AAA expansion (p=0.042) and rupture or repair (HR 2.49, 95% CI1.07, 5.78; p=0.034), even when adjusted for age, sex, body mass index, systolic blood pressure, current smoking and, crucially, aneurysm diameter. Conclusion These are the largest USPIO MRI and PET-CT studies in AAA disease to date and the first to investigate 18F-NaF. Both USPIO MRI and 18F-NaF PET-CT are able to predict AAA expansion and the composite of rupture and repair, with 18F-NaF PETCT emerging as the first imaging biomarker that independently predicts expansion and AAA events, even after adjustment for aneurysm diameter. This represents an exciting new predictor of disease progression that adds incremental value to standard clinical assessments. Feasibility and randomised clinical trials are now required to assess the potential of this technique to change the management and outcome of patients with AAA

Risk stratification in atherosclerotic cartoid stenosis

Introduction: Key trials and a Cochrane systematic review in asymptomatic carotid stenosis have highlighted the need to identify a high-risk subgroup of patients with carotid stenosis who may benefit from intervention. Traditionally, this risk stratification has considered structural imaging and clinical factors. However, using only these approaches, still a significant number of patients are missed. Biological attributes are acknowledged as key determinants of thrombo-embolic events. Functional and hybrid structural-functional imaging, and circulating biomarkers allow exploration of plaque biology non-invasively, in vivo. The importance of innate immunity in atherosclerosis is now established, with a recent interest in macrophage phenotypic polarisation in atherosclerosis supported by in vitro and experimental data, with the hypothesis of an M1 macrophage predominance associated with unstable plaques. The emergence of systems biology has been seen to facilitate understanding of biological pathways and generate hypotheses, although the utility of this approach for the examination of human atherosclerosis tissue has not been fully explored. Aims: (i) To employ functional imaging to probe carotid atherosclerosis in vivo; (ii) to assess the plaque microenvironment in determination of the balance of macrophage populations in unstable compared with stable atherosclerosis; (iii) to investigate whether late phase (LP-) contrast enhanced ultrasound (CEUS) reflects plaque biological features; (iv) to examine the utility of systems biology techniques in distinguishing symptomatic from asymptomatic carotid atherosclerosis tissue, and in hypothesis generation; and (v) to evaluate a putative biomarker for carotid atherosclerosis and plaque vulnerability. Methods: Patients with carotid stenosis, both symptomatic and asymptomatic, have undergone systematic collection of data, fresh carotid endarterectomy (CEA) specimens, and plasma. Thirty-two patients with 36 carotid stenoses underwent 11C-PK11195 PET/CT. Thirty-seven patients had dynamic (D-) and LP-CEUS carotid imaging. CEA specimens were assessed by immunohistochemical techniques, as well as atheroma cell culture with supernatant multi-analyte profiling (MAP). MAP data was subject to Ingenuity Pathway Analysis. CEA specimens were further examined using systems biology methodologies: transcriptomics with Affymetrix Human Exon 1.0 ST arrays; proteomics and lipidomics by liquid chromatography (LC) coupled to tandem triple quadrupole mass spectrometry (MS); and metabolite profiling by nuclear magnetic resonance and LC-MS. Furthermore, venous and arterial plasma was quantified for the lysozyme, a putative biomarker in carotid atherosclerosis. Results: 11C-PK11195 PET allowed the non-invasive quantification of intraplaque inflammation in patients with carotid stenoses and, when combined with CTA, provided an integrated assessment of plaque structure, composition and biological activity. 11C-PK11195 PET/CT distinguished between recently symptomatic vulnerable plaques and asymptomatic plaques with a high positive predictive value. D-CEUS and LP-CEUS (at a cut-off of zero) was able to distinguish symptomatic and asymptomatic plaques. Atheroma cell culture and supernatant MAP revealed that symptomatic human atherosclerotic carotid disease is associated with a cytokine and chemokine pattern consistent with the predominance of pro-inflammatory M1-type macrophage polarisation. Furthermore, IFNγ signatures are observed, including the novel finding of CCL20 with its significant elevation in symptomatic atherosclerosis. MAP of supernatants from patients who had undergone ipsilateral carotid LP-CEUS revealed significantly higher levels of IL6, MMP1 and MMP3, as well as greater CD68 and CD31 immunopositivity, in those with high (≥0) compared with low (<0) LP-CEUS signal. This suggests that LP-CEUS was able to reflect plaque biology. Transcriptomic analysis was able to clearly separate stenosing plaque and intimal thickening, as well as unstable and stable atherosclerosis, finding differential expression and alternative splicing of interferon regulatory factor 5 between stenosing plaque and intimal thickening. Proteomic analysis of the salt extract fraction from carotid atherosclerotic plaques identified 2,470 proteins implicated in 33 bio-molecular functions and having their origins previously described in 14 different cellular compartments. There were 159 proteins which, based upon the number of assigned spectra, were significantly different between symptomatic and asymptomatic atherosclerosis. Through lipidomic analysis, 150 lipid species from 9 different classes were identified, of which 24 were exclusive to atherosclerotic plaques. A comparison of 28 carotid endarterectomy specimens revealed differential lipid signatures of symptomatic compared with asymptomatic lesions, as well as stable and unstable plaque areas. Similarly, LC-MS metabolite profiling of organic plaque extract was able to separate symptomatic from asymptomatic atherosclerosis. Arterial and venous plasma lysozyme levels were seen to distinguish individuals with carotid atherosclerosis from matched control subjects. Furthermore, arterial plasma lysozyme levels were significantly higher in patients with symptomatic than asymptomatic carotid stenosis. Conclusions: These findings support the use of hybrid structural-functional imaging, and the utility and use of a systems biology approach in identifying significantly different and biologically relevant variations in atherosclerosis tissue, and in hypothesis generation for further study. The data presented concurs with recent reports in the literature linking the lipidic/organic component of atherosclerosis with the generation of a pro-inflammatory plaque microenvironment prone to lesion development, instability and the complications thereof. The importance of innate immunity has been highlighted with the demonstration of a predominance of M1 macrophage polarisation and evidence of Th17/IL17 signalling in unstable atherosclerosis. It is hoped that this work will contribute to the ongoing refinement of multi-factorial risk stratification in carotid atherosclerosis

A retrospective study of renal dysfunction in acute stroke: incidence, impact and outcomes

Stroke is a leading cause of death and neurological disability worldwide. Chronic kidney disease (CKD) is associated with an increased risk of stroke. Conversely, CKD confers worse outcomes following a stroke, with the highest mortality seen in end-stage renal disease. In comparison, the relationship between AKI and stroke is not well described, with a lack of UK data. In this single-centre, retrospective observational study of hospitalisations with acute stroke, I sought to determine the incidence of renal dysfunction and its impact on outcomes. AKI incidence was determined using preadmission serum creatinine (SCr) as ‘baseline’ renal function, compared with 4 surrogate methods. AKI was common, with an overall incidence of 20%, and was associated with increased 30-day and 1-year mortality using all AKI methods. Admission SCr most closely agreed with preadmission SCr but all surrogate methods exhibited bi-directional misclassification of AKI. CKD prevalence was high (over 30%) and was associated with increased mortality in univariable analyses. CKD patients underwent fewer imaging modalities and thrombectomy, possibly suggesting the presence of ‘renalism’. Contrast exposure was not found to be a risk factor for AKI. Vascular calcification and carotid artery disease were univariably associated with CKD. Multi-centre studies are needed to confirm the findings

Neuroimaging - Clinical Applications

Modern neuroimaging tools allow unprecedented opportunities for understanding brain neuroanatomy and function in health and disease. Each available technique carries with it a particular balance of strengths and limitations, such that converging evidence based on multiple methods provides the most powerful approach for advancing our knowledge in the fields of clinical and cognitive neuroscience. The scope of this book is not to provide a comprehensive overview of methods and their clinical applications but to provide a "snapshot" of current approaches using well established and newly emerging techniques