The literature on stroke reveals an increasing interest in the role played by social and emotional factors in rehabilitation after stroke. A comprehensive literature survey shows profiles of spontaneous recovery, the significance of a team approach to rehabilitation, patterns of prognostic significance for long-term recovery and adaptation and formulations of rehabilitation models for the Western world. The importance of depression as a major factor in demotivation to participate in rehabilitation and achieve long-term quality of life post-stroke emerges. From the literature survey a research design was formulated for the ecological study of a sample of 51 stroke patients at Ga-Rankuwa Hospital near Pretoria. The questionnaire was structured according to the Synchronous Systems Model, and data gathered from the biological, personal and environmental spheres of patients. Data was collected by a multidisciplinary team at three assessment times, three days, two weeks and three months post-stroke. These corresponded to the acute physical phase of stroke, the end of the hospitalisation period, and an assessment of patients once they had been discharged back into the community. Descriptive statistics were obtained on all variables and principle axis factor analysis was performed to verify the factorial structure of the tests. In order to establish whether group scores changed between assessments, t-tests for dependent measures were applied. Pearson Product Moment correlations were computed for the purpose of establishing relationships between variables. The results revealed dramatically differing biographical characteristics of the sample of stroke patients both premorbidly and at three months after the stroke. Significant recovery profiles emerged in both the physical and neuropsychological spheres at both the 14 day and 3 month assessments. Depression and the functioning at home and at work social sphere of role emerged as profiles of deterioration. At 14 days, depression was related to physical and cerebral functioning. This changed at three months, with depression also being significantly related to aspects of social functioning. On the basis of these results, depression after stroke was conceptualised as a severance of relational connectedness in the social ecological functioning of stroke patients. An ecological approach to rehabilitation is proposed that would seek to reframe the identity of stroke patients and establish relational connectedness post-stroke. / Psychology / D. Litt. et Phil. (Psychology)

Cerebral ischaemia, or stroke, is a leading cause of death and disability worldwide. Ischaemic stroke, as a result of arterial occlusion, and subarachnoid haemorrhage (SAH), as a consequence of arterial rupture in the subarachnoid space, are major subtypes of stroke. Treatment options for both are limited, and many therapeutic strategies have failed. In ischaemic stroke, lack of evidence of brain penetration of treatments has been cited as a major weakness and contributing factor to failed clinical trials. In SAH, animal models do not always mimic key pathophysiological hallmarks of the disease, hindering development of new therapeutics. Inflammation is strongly associated with brain injury after cerebral ischaemia and inhibition of the pro-inflammatory cytokine interleukin-1 (IL-1) represents apossible therapeutic target. Therefore, the key objectives of this thesis were; (1) to improve preclinical data on a promising stroke treatment, interleukin-1 receptor antagonist (IL-1Ra), by investigating its pharmacokinetic profile and brain penetration in a rat model of ischaemic stroke, (2) to investigate the endovascular perforation model of SAH in rat, as a tool for the investigation of neuroprotectants, and (3) to examine the role of the inflammatory response in the SAH model and the effects of IL-1Ra. The neuroprotective effect, pharmacokinetic profile and brain penetration of IL-1Ra were assessed after a single subcutaneous (s.c.) dose (100mg/kg) in rats, after transient (90 min) middle cerebral artery occlusion (MCAo). A single s.c. dose of IL-1Ra reduced neuronal damage, resulted in sustained, high concentrations of IL-1Ra in plasma and cerebrospinal fluid and also penetrated brain tissue exclusively in areas of blood brain-barrier (BBB) breakdown. An endovascular perforation model of SAH in rat was investigated and produced widespread multifocal infarcts. In this model, administration of IL-1Ra (s.c.) reduced BBB breakdown, which correlated with injury at 48 h. IL-1_ was expressed in the brain early after SAH in areas associated with haem oxygenase-1 (HO-1) expression, indicating the presence of free haem. Stimulation of primary mouse mixed glial cells in vitro with haem induced expression and release of IL-1 alpha but not IL-1 beta. These data, after MCAo in rat, are the first to show that a single s.c. dose of IL-1Ra rapidly reaches salvageable brain tissue and is neuroprotective. This allows confidence that IL-1Ra is able to confer its protective actions both peripherally and centrally. After experimental SAH, we suggest that haem, a breakdown product of haemoglobin, released from lysed red blood cells in the subarachnoid space, acts as a danger associated molecular pattern (DAMP) driving IL-1- dependent inflammation. These data provide new insights into inflammation after SAH-induced brain injury and suggest IL-1Ra as a candidate treatment for the disease. Overall, these findings strengthen preclinical data supporting IL-1Ra as a neuroprotective therapy for ischaemic stroke, and identify SAH as a new indication for treatment with IL-1Ra.

Progesterone has been shown to be neuroprotective in a number of central nervous system injury models, including cerebral ischaemia. There is still a lack of understanding behind progesterone’s neuropotective properties, and the purpose of this project is to clarify some of these issues. Osmotic mini-pump infusion was hypothesised to more effective in delivering progesterone to the target organ of the brain, when compared to a bolus intraperitoneal injection. Progesterone pharmacokinetic profiles were compared between different dosing regimes. Intraperitoneal progesterone injection had a short half-life in both plasma and brain, while osmotic mini-pumps delivered higher concentrations of progesterone in plasma and particularly in brain, over a longer period, which supports the hypothesis. It was hypothesised that progesterone will reduce NO production and cell death in in vitro. Progesterone reduced nitric oxide production after challenging microglia with LPS, which supports our hypothesis and the nuclear progesterone receptor was found not to have a major role in nitric oxide attenuation. Neither of the microglial cell lines, BV-2 and HAPI cells produced elevations in NO formation in ischaemic conditions. The in vitro oxygen and glucose deprived model of ischaemia, reduced viability in both microglial and neuronal cells. Also, high pharmacological concentrations of progesterone exacerbated ischaemic injury, which does not support the hypothesis of progesterone in reducing cell death. Progesterone administration, via osmotic mini-pump infusion, was hypothesised to have a better outcome compared to vehicle treatment. After the onset of experimental stroke, progesterone delivery via osmotic mini-pump with loading dose was found to be beneficial in terms of neurological deficit score in adult male mice, which supports the hypothesis. Also, we hypothesise that co-morbidity can affect the efficacy of progesterone treatment in outcomes. Aged animals have an increased sensitivity to experimentally induce stroke and did not display, in the outcomes measured, any benefit from progesterone treatment. NOD/ShiLtJ mice had severe symptoms, resulting in high mortality after surgery and are not recommended as a model of diabetes for experimental stroke. Hypertensive BPH/2 mice are a potential hypertensive model and had better functional outcomes after treatment with intraperitoneally administered progesterone, compared to non-treated hypertensive animals in our small preliminary study. This supports our hypothesis that co-morbidity can affect the efficacy of progesterone treatment in outcomes. The gold-standard for assessing intervention effects across studies within and between subgroups is to use meta-analysis based on individual animal data. We hypothesise meta-analysis would reveal progesterone to reduce lesion volume, but also discover other effects in different subgroups of animals. Progesterone significantly reduced lesion volume, it also appeared to increase the incidence of death following experimental stroke. Furthermore, this negative effect appears to be particularly apparent in young ovariectomised female animals. These findings support the hypothesis that progesterone reduces lesion volume and progesterone having other effects in different subgroups. This investigation has clarified some issues and expanded our understanding on the neuroprotective properties of progesterone. However, these findings indicate further investigation is still required before progesterone can be considered for use in clinical trials as a neuroprotectant in stroke.

Apolipoprotein E (apoE denotes protein: APOE denotes gene) is a lipid-transport protein abundantly expressed in the brain and strongly upregulated after acute brain injury. The APOE e4 allele is the major genetic risk factor for Alzheimer’s disease (AD) and has been associated with poorer outcome after various types of acute brain injury, including traumatic brain injury and subarachnoid haemorrhage. However, the role of APOE genotype in focal ischaemic stroke is less clear. The mechanism(s) by which APOE genotype may modulate outcome after acute brain injury are also unclear at present. Accordingly, the studies described in this thesis were undertaken to further address these issues. 1. Endocytic pathway alterations in human temporal lobe after global cerebral ischaemia and association with APOE genotype. 2. Characterisation and validation of the intraluminal filament model of focal cerebral ischaemia in C57BI/6J mice. 3. Association between APOE genotype and differences in outcome and endocytic pathway alterations after focal cerebral ischaemia in mice. 4. Adenovirus-mediated gene transfer of APOE e3 markedly reduces ischaemic brain damage after focal cerebral ischaemia in mice. The data presented in this thesis indicate an important role for APOE genotype in modulating outcome after ischaemic brain injury, further highlighting the favourable effects associated with the APOE e3 allele. APOE genotype-dependent alterations in the endocytic pathway are mechanisms which could contribute to differences in outcome. These data also highlight the neuroprotective effects achieved by manipulating apoE levels to promote the beneficial effects of apoE3. An apoE-based therapeutic strategy may be a potential approach for treatment of ischaemic brain injury in humans.

National guidelines for stroke recommend that all patients entering rehabilitation are screened for mood disturbance using a validated measure. The first half of this thesis presents a literature review of 25 self-report screening measures for the detection of post-stroke distress. A total of 26 studies were identified as meeting the search criteria. Fifteen self-report measures met recommended levels of sensitivity (≥0.80) and specificity (≥0.60) when screening for post-stroke depression. The Hospital Anxiety and Depression Scale (HADS) was the only measure to meet recommended levels of accuracy for post-stroke anxiety. At the commencement of this thesis, the Distress Thermometer (DT) had not been validated among stroke survivors despite being recommended by NICE (2009). The study presented in the second half of this thesis investigates the diagnostic accuracy and clinical utility of the DT and associated Problem List (PL), the Brief Assessment Schedule Cards (BASDEC), and the Yale. Relative to the HADS, the area under the curve (AUC) for the DT was significantly greater than an AUC of 0.50. Cut-off scores of at least 4 and 5 on the DT met recommended levels of sensitivity and specificity when screening for post-stroke depression and anxiety. The accuracy of the BASDEC and Yale was non-significant. Due to a small sample size, these results should be taken with caution. However, this study provides preliminary evidence to support the use of the DT and PL as a holistic and person-centred screening tool for the prevention and recognition of post-stroke distress.

Background: Hyperglycaemia is common in acute stroke and is associated with a poor outcome. Underlying aetiology and mechanism of action is poorly understood. Management remains uncertain. Methods: We undertook a randomised placebo controlled trial to assess the effect of GKI (Glucose-Potassium-Insulin) versus placebo on lesion volume progression and cerebral lactate levels using magnetic resonance imaging (MRI) and spectroscopy (MRS). An observational study of the capillary blood glucose within 48 hours of stroke onset was performed to define the temporal profile of glucose, with a subset followed prospectively to determine the prevalence of abnormal glucose metabolism in patients with stress hyperglycaemia. The association between insular cortex involvement and hyperglycaemia was determined by analysing MRI data sets from two randomised trials. Stroke unit practice for the management of glucose was assessed in a review of the stroke unit trialists’ collaboration data set. Results: • GKI infusion failed to attenuate infarct growth in patients with moderate hyperglycaemia within 24 hours of acute ischaemic stroke. A trend towards attenuation of increased lactate concentration was evident in the GKI treatment arm. Exploratory analyses raised the possibility that GKI may be harmful in patients with persistent arterial occlusion. • Over the 48hour monitoring period 75% of patients developed Hyperglycaemia. Stroke severity was not predictive of admission hyperglycaemia whereas glycosylated haemoglobin was (OR 2.97; 95%CI 1.84-4.78; p<0.001). 50% of patients screened were found to have abnormal glucose metabolism at follow-up. • Insular cortex involvment on MRI was not predictive of admission hyperglycaemia. • Testing for blood glucose concentration in stroke units was infrequent. Of the minority of units that had a protocol in place, the threshold for intervention with insulin was >10mmol/l. Conclusion: We found no evidence that GKI infusion attenuated infarct growth in patients with mild hyperglycaemia following acute ischaemic stroke. In post-hoc analysis the possibility that GKI infusion may be harmful in patients with total occlusion suggests an effect dependent on recanalisation status. A non-significant trend towards attenuation of increased lactate concentration was evident. Stroke severity was not found to be a predictor of post stroke hyperglycaemia. Underlying dysglycaemia was common in non-diabetic patients manifesting hyperglycaemia within 48hours of stroke ictus. Screening of high risk patients with oral glucose tolerance testing is justified and provides a potential opportunity for secondary prevention. Insular cortex involvement did not independently predict hyperglycaemia in acute stroke. Current management of hyperglycaemia is guided by consensus guidelines with little evidence base. Stroke unit practice varies with little change across stroke units over the years.

In the UK, stroke is the third most common cause of death after heart disease and cancer. Importantly, most strokes are not fatal meaning stroke is the leading cause of adult disability, with one third of survivors still functionally dependent after one year. Post stroke hyperglycaemia (PSH) predicts poor outcome independent of age, stroke type or severity and occurs in over 60% of patients without a diagnosis of diabetes and in more than 90% of diabetic patients. Around 56% of hyperglycaemic acute stroke patients are subsequently diagnosed with insulin resistance, manifesting as impaired glucose tolerance, impaired fasting glucose or the “metabolic syndrome.” Clinical guidelines recommend routine blood glucose monitoring, and intervention with insulin for hyperglycaemia. However, there is currently no clinical evidence of benefit from insulin use, and recent findings raise concerns about the safety of insulin use in predominantly non-diabetic acute stroke populations. Hyperglycaemia in animal models of cerebral ischaemia is associated with increased infarct size. However, a recent systematic review highlighted uncertainties about the relevance of many previously published preclinical studies to the typical clinical picture of PSH with respect to whether clinically relevant elevations in blood glucose exacerbate ischaemic brain damage in models other than those of type I diabetes, or whether responses are different in animals with and without features of metabolic syndrome. The aim of this thesis was to investigate the differential effects of clinically relevant hyperglycaemia on the acute evolution of damage in animals with and without features of the metabolic syndrome and to investigate the potential mechanistic role of oxidative stress in glucose mediated ischaemic damage. I hypothesised that hyperglycaemia would exacerbate acute lesion evolution and final infarct volume in animals with and without features of the metabolic syndrome and that oxidative stress mechanisms would be involved. Developing suitable animal models for investigating post stroke hyperglycaemia and metabolic syndrome The first aim of this thesis was to develop a series of animal models for the investigation of clinically relevant PSH, metabolic syndrome in the presence and absence of PSH, and an appropriate focal cerebral ischaemia model. A previously described model of metabolic syndrome using the spontaneously hypertensive stroke-prone rat (SHRSP) fed on a 60% fructose diet for two weeks was set up in the laboratory and glucose tolerance, adiposity, plasma insulin, plasma triglycerides, cholesterol were measured and compared to the SHRSP reference strain, the Wistar-Kyoto (WKY). Fructose fed-SHRSP rats exhibited glucose intolerance, hypertriglyceridaemia, hyperinsulinaemia, increase adiposity and reduced HDL cholesterol compared to the WKY controls. This suitably modelled the features of the metabolic syndrome. Clinically relevant levels of PSH were achieved using a bolus intraperitoneal injection of 15% glucose (10ml/kg) 10 minutes prior to permanent middle cerebral artery occlusion (MCAO). Two methods of MCAO were compared; intraluminal filament (ILF) model and a distal diathermy MCAO model. The ILF method produced expansive lesions incorporating the majority of the ipsilateral hemisphere whereas the distal diathermy occlusion produced smaller cortical infarcts which were shown to produce a large volume of penumbra. The diathermy model was selected for all future experiments. Investigating the effects of hyperglycaemia on acute lesion growth after focal cerebral ischaemia in rats with and without components of the metabolic syndrome The second aim of this thesis was to determine the effects of hyperglycaemia on acute ischaemic lesion evolution and infarct volume in rats with and without features of the metabolic syndrome. Hyperglycaemia was induced in WKY or fructose-fed SHRSP rats 10 minutes prior to permanent MCAO. Magnetic resonance imaging (MRI) was used to quantify the expansion of the lesion using ADC maps calculated from diffusion weighted imaging (DWI) carried out over the first 4 hours after MCAO. Acute hyperglycaemia, at clinically relevant levels, exacerbated early ischaemic damage in both normal and metabolic syndrome rats. Hyperglycaemia worsened infarct volume at 24h in normal but not metabolic syndrome rats. These data suggest that management of hyperglycaemia may be most beneficial in the absence of an underlying dysglycaemia. Using a marker of oxidative stress to probe the tissue sections suggested that perhaps hyperglycaemic effects were mediated through oxidative stress mechanisms. Investigation of oxidative stress mechanisms in hyperglycaemia-dependant ischaemic damage. Based on the potential involvement of oxidative stress mechanisms in hyperglycaemia-associated ischaemic damage this was investigated in a proof of concept study using pre-treatment with a SOD/catalase mimetic, EUK-134. The rationale for this study was based on the involvement of increased superoxide production in chronic hyperglycaemia and diabetic complications. Sprague-Dawley rats received the same dose of glucose previously used 10 minutes prior to MCAO by distal diathermy. EUK-134 (2.5mg/kg) or vehicle (saline) was administered 20 minutes prior to MCAO. Infarct volume, calculated from RARE T2-weighted MR images and neurological scoring was measured 24 hours post MCAO. Arterial superoxide levels were measured along with the lipid peroxidation marker malondialdehyde in cerebral tissue. Hyperglycaemia increased infarct volume although no effect of high blood glucose on tissue lipid peroxidation was observed. EUK-134 failed to reduce infarct volume. EUK-134 reduced lipid peroxidation in ipsilateral cortex of normoglycaemic rats but not in hyperglycaemic rats. Thus, hyperglycaemia associated damage was not reduced by attenuating oxidative stress. Generating a threshold for quantitative T2 maps Using the data generated in the previous Chapter a study was carried out to determine if improvements could be made in the analysis of MRI data in pre-clinical rat studies. RARE T2-weighted images, where T2 relaxation time denotes contrast, have been used for infarct quantification 24 hours post MCAO. Generating a lesion volume from these images requires manual tracing of the visually defined infarct from the images. A degree of error is associated with this methodology either as inter-investigator reproducibility or in repeats conducted by the same investigator. The RARE T2 images are not quantifiable as arbitrary values denote contrast and this prevents direct comparison of T2 relaxation times for a particular structure between subjects. Using a MSME T2 sequence, quantitative T2 maps can be generated which calculate absolute T2 relaxation times for each voxel. Using quantitative maps an attempt was made to establish a threshold to discriminate between normal and ischaemically damaged tissue in animals that had undergone MCAO using the distal diathermy model. This threshold aimed to minimise inter- or intra-investigator error. A T2 threshold of abnormality for the quantitative T2 maps was established from the RARE T2 weighted images. Tissue that had a T2 relaxation time of 76 ms or more was deemed to be infarct according to the derived threshold. The suitability of the threshold was then assessed in a separate cohort and was found to produce similar infarct volumes to the manual delineation of RARE T2-weighted images. The 76ms threshold was also applied to animals that had undergone MCAO using the intraluminal filament model. However, the calculated threshold was not suitable for the intraluminal filament model due to interferences from cerebrospinal fluid signal. A more selective method for determining infarct volume in animals subjected to intraluminal filament MCAO needs to be established.

Following stroke, a patient may fail to report touch administered by another person but claim that s/he feels touch when it is self-administered. In Part One, the self-touch rubber hand paradigm was used to investigate different explanations for this phenomenon, termed self-touch enhancement. The most important finding was that patients reported touch based on feeling rather than by using proprioceptive information. Some patients have residual sensation that could be targeted in sensory rehabilitation. Part Two is a systematic investigation of the illusion of self-touch conducted with neurologically healthy participants. Participants used the right hand to administer touch to a prosthetic hand while the left (receptive) hand, positioned 15 cm from the prosthetic hand, received Examiner-administered touch. Proprioceptively perceived position of the administering and receptive hand was measured. Most participants experienced the single event of self-touch at the location of the receptive hand. Previous investigations have relied on measurement of only one hand and have concluded that participants experience self-touch at the location of the prosthetic hand. Our findings have implications for the role of ownership in this illusion. There is also a series of experiments in Part Two which test four potential constraints on the illusion of self-touch – violated expectations about the object that is administering touch, increased distance between the hands, alignment mismatch, and anatomical implausibility. For example, one study uses a novel paradigm to demonstrate that, although the subjective intensity of the illusion of self-touch is diminished by anatomical implausibility, most participants report the impossible experience of touching their left elbow with their own left index finger. Taken together, these experiments highlight the malleability of body representation, and provide a comprehensive framework for understanding the illusion of self-touch.

En France, on estime à 150 000 le nombre d’accident vasculaire cérébraux par an. Avec un coût de 8,3 milliards d’euros, c’est devenu une problématique de santé publique. Les dépenses sont, entre autres, lié aux séquelles, car 43,2 % sortent d’hospitalisation en étant dépendant physiquement.Ces séquelles vont séquelle vont être à l’origine de limitations, provoquant un cercle vicieux d’inactivité, et amenant un déconditionnement. Pour lutter contre ce déconditionnement, il semble pertinent de respecter les recommandations des sociétés savantes en termes d’activité physique (AP). Néanmoins en retour à domicile, très peu de patients atteignent le minimum d’AP requis. L’éducation thérapeutique du patient (ETP) semble être une option pertinente pour remédier à ce problème.Ce travail de recherche s’est donc intéressé à l’effet d’un programme ETP à l’AP chez des patients en phase subaigue. Le programme était effectué au domicile des patients, et était composé de visites à domicile et d’appels téléphoniques. L’AP était contrôlée grâce à un actimètre et des livrets de suivi.Après les 6 mois du programme, il ressort une augmentation du périmètre de marche, mais pas d’impact sur la force des membres inférieurs, la composition corporelle, la fatigue et la dépression.Les principales caractéristiques des patients volontaires à un programme ETP à l’AP étaient la persistance de la fatigue, la dépression, peu de séquelles neuro-motrices physiques et un fort soutien de l’entourage. Enfin, au cours des 6 mois de suivi, les patients ont maintenu des durées d’AP qui étaient au-dessus des recommandations en termes d’AP. / In France, we estimate at 150,000 the number of cerebral vascular accident per year. With a cost of 8.3 billion euros, it has become a public health issue. The expenses are, among others, related to legacy, as 43.2% leave hospital being physically dependent.These effects will aftereffect will cause limitations, causing a vicious cycle of inactivity and deconditioning bringing. To fight against this deconditioning, it seems appropriate to follow the recommendations of learned societies in terms of physical activity (PA). Yet back home, very few patients reach the minimum required AP. Therapeutic patient education (TPE) seems to be a relevant option to address this problem.This research is therefore interested in the effect of ETP to the AP in patients with subacute phase. The program was conducted in patients' homes, and consisted of home visits and phone calls. The PA was controlled through a actimeter and monitoring booklets.After 6 months of the program, it appears an increase in the walking distance, but no impact on the strength of lower limbs, body composition, fatigue and depression.The main characteristics of volunteer patients to ETP to the PA were persistent fatigue, depression, few physical neuro-motor sequelae and strong support from the entourage.Finally, during the 6 months follow-up, patients maintained durations of AP which were above recommendations in terms of AP.

Clinical manipulation of angiogenesis (the formation of new blood vessels from pre-existing vasculature) is of interest to treat diseases such as cancer and ischemic tissue where it is not properly regulated. Several treatments targeting vascular endothelial growth factor (VEGF) and its receptors - which are abundant at sites of angiogenesis - are currently in use to treat various types of cancer, however they have severe vascular side effects. Conversely, VEGF has been used clinically to promote angiogenesis to treat ischemic tissue. However, despite encouraging data from pre-clinical models, trials in humans have been disappointing. For further therapies to be developed, more information on how VEGF interacts with its receptors is required. Heparan sulfate (HS) is a ubiquitous glycosaminoglycan involved in a number of physiological processes including angiogenesis. HS facilitates the interaction of VEGF with its receptors, which is crucial for angiogenesis. Modification of this interaction via synthetic mimetics of HS may allow clinical intervention of angiogenesis. The current investigation aims first, to clarify the requirement for the interaction between VEGF and HS in angiogenesis; second to characterise the structure of HS that binds to VEGF so that mimetics can be developed; and third, to determine the effect of HS mimetics on angiogenesis in vivo. To determine the requirement for VEGF/HS interaction in angiogenesis, several mutants of VEGF165 that had lower affinities for HS were assayed for their ability to induce ectopic angiogenesis in the subintestinal baskets of zebrafish embryos. Wild type VEGF165 induced a 200-250% increase in ectopic vessels, which was matched only by a control mutant. Other mutants did not induce ectopic vessels, suggesting that this interaction is required for angiogenesis. To characterise the structure of HS that binds to VEGF, various HS mimetics were assayed against heparin in a VEGF competition assay using Biacore. Of these, the strongest inhibition (IC¬50 =~16nM) was with 2O10, an oligosaccharide that consisted of two highly sulfated octasaccharide domains (NS domains) that flanked an unsulfated dodecasaccharide region. To determine the type of sulfation required for this interaction, HS fragments were assayed for interaction with VEGF165 using the filter binding assay, and analysed by HPLC which indicated 6-O sulfation may be preferential for VEGF binding to HS.To investigate the ability of HS to affect angiogenesis, the effects of HS mimetics on zebrafish embryo subintestinal baskets were measured. The most interesting of these was with 2O10, which had a biphasic response whereby low doses (3ng) increased basket vasculature by 30% and high doses (30ng) decreased the endogenous vessels by 20%. As 2O10 had a high affinity for VEGF, its effects on the vasculature may be due to interaction with endogenous VEGF, which would indicate that HS mimetics can be used to control angiogenesis by modification of growth factor signalling. The investigation concludes that the interaction between VEGF and HS is critical for angiogenesis, and that this can be modulated by the application of HS mimetics that bind strongly to VEGF.