The hemocompatibility of crystalline Silicon Carbide (SiC), in its cubic form (i.e., 3C-SiC), has been evaluated and compared to Silicon (Si), the leading material in biosensing applications. Silicon carbide (SiC) is a hard, chemically robust material, very well suited for harsh environment applications, and has been suggested to have very good biocompatibility. Additionally, SiC in its amorphous form, has been used as a coating for medical implantable devices such as bone prosthetics and cardiovascular stents. However, assessment of single crystal 3C-SiC for cardiovascular applications has not been reported. In this research we have studied the interactions of single crystal 3C-SiC with platelets and human microvascular endothelial cell (HMVEC) to assess the degree of hemocompatibility of 3C-SiC. The more hemocompatible a material is, the less platelet adhesion would be expected. Using fluorescence microscopy higher platelet adhesion was statistically observed on Si than on SiC. In addition 3C-SiC surfaces showed less platelet reactivity, measured by the degree of platelet adhesion, aggregation and activation, with mostly circular morphology of adhered platelets while Si showed an elevated presence of non-activated (Circular) platelet clumps. Additionally, HMVEC proliferation assessment suggest that 3C-SiC performs comparably to high attachment culture wells with enhanced proliferation, without affecting cell morphology. These results suggest that 3C-SiC is a promising candidate for applications in the blood stream due to its low thrombogenic characteristics and good hemocompatibility.

No description available.

published_or_final_version / abstract / Medicine / Master / Master of Philosophy

published_or_final_version / Medical Sciences / Master / Master of Medical Sciences

A substantial number of patients undergoing cardiac surgery are on dual anti-platelet treatment with clopidogrel and aspirin. A disadvantage with this treatment is increased risk of bleeding. Bleeding is a complication of major concern associated with adverse outcome for the patient and increased hospital resource utilization. Great variability in individual response to clopidogrel has been reported. If in vitro measurements of platelet reactivity would correlate with clinical bleeding parameters, potential bleeders could be identified preoperatively. The aims of this thesis were: (1) to describe the degree of pre-operative platelet inhibition in patients scheduled for primary isolated coronary artery bypass graft surgery; (2) to prospectively investigate whether the pre-operative platelet inhibition correlated with intra- and postoperative bleeding and transfusion requirements; and (3) to test the ability of clinically relevant risk factors to predict re-exploration for bleeding. (4) In addition, a cost analysis was performed on patients re-explored for bleeding, to analyse the magnitude of added resource utilization and costs. Based on this, a cost model of prophylactic treatment with haemostatic drugs was calculated. Platelet function tests investigated were: (1) flow cytometry, (2) VASP, (3) VerifyNowSystem, (4) PlateletMapping (a modified TEG), and (5) PFA-100. Clinical risk factors for re-exploration and the influence of antiplatelet and antifibrinolytic therapy were evaluated in a retrospective analysis. Cost analysis at three cardiothoracic centres was performed in a case-control study. In conclusion, there was no clinically useful correlation between preoperative assessment of platelet inhibition and blood loss or transfusion requirements during coronary artery bypass surgery. Furthermore, there was only modest agreement between the methods evaluating ADP-receptor blockade. Pre-operative treatment with the P2Y12-receptor inhibitor clopidogrel was an essential risk factor for re-exploration due to bleeding. Except for clopidogrel, no strong clinical factor to predict the risk of re-exploration was identified. The resource utilisation costs were 47% higher in patients requiring re-exploration due to bleeding than in those not requiring re-exploration. Prolonged stay in the ICU and recovery ward accounted for half of the added cost, a third was due to the costs of surgery, one fifth due to increased cost of transfusions, and <2% was due to haemostatic drug treatment.

1. Ištyrus homocisteino koncentraciją kraujo serume ir trombocitų agregaciją kraujo plazmoje su pagrindiniais natūriniais agregantais (adenozino difosfatas, adrenalinas, kolagenas) sergančiųjų ūminiais galvos smegenų kraujotakos sutrikimais, nustatyta koreliacija tarp:  padidėjusios homocisteino koncentracijos ir insulto su dideliu neurologiniu deficitu, išeminės širdies ligos, kreatinino ir C-reaktyviojo baltymo koncentracijų;  vidinės miego arterijos stenozės laipsnio ir homocisteino koncentracijos kraujo serume;  homocisteino koncentracijos kraujo serume ir amžiaus;  padidintos homocisteino koncentracijos ir pasikartojančio insulto, todėl homocisteino koncentracijos lygis gali būti panaudotas kaip prognostinis rodiklis naujam ar kartotiniam ūminiam galvos smegenų kraujotakos sutrikimui išsivystyti. 2. Trombocitų agregacija turtingoje trombocitais plazmoje ūminio išeminio galvos smegenų kraujotakos sutrikimo periode statistiškai patikimai padidėja insultu sergančiųjų grupėje, nepriklausomai nuo susirgimo sunkumo. 3. Profilaktinės aspirino dozės hiperhomocisteinemijos fone dalį ligonių neapsaugo nuo išeminio insulto išsivystymo. Aspirino vartojimas turi veiksmingesnį antiagregacinį poveikį moterims, nei vyrams: moterų plazmoje trombocitų agregacijos intensyvumas buvo žemesnis su visais agonistais, o vyrų trombocitai, atvirkščiai, į juos reagavo viršnorminiu atsaku. 4. Išeminiu insultu sirgusių ligonių kraujyje dažniau randami policitemija, padidėjęs leukocitų... [toliau žr. visą tekstą] / 1. When examining homocysteine concentration in blood serum and platelet aggregation in plasma using nature aggregants (adenosine diphosphate, adrenalin, collagen) in patients with acute cerebrovascular disorders, the positive corelation was found between the following parameters:  the elevated amount of homocysteine and stroke with the high neurological deficit, ischemic heart disease, creatinin and C-reactive protein concentrations;  the degree of internal carotid artery stenosis and homocysteine concentration in blood serum;  homocysteine concentration in blood serum and age;  elevation of homocysteine concentration and recurrent stroke, thus, homocysteine concentration may be a predisposing indicator to the development of new or recurrent acute cerebrovascular disorder. 2. Platelet aggreagation in platelet-rich plasma during the period of acute cerebrovascular disorder statistically significantly increases in stroke group independent upon severity of illness. 3. Preventive aspirin doses in the light of hyperhomocysteinemia do not protect a certain part of patients from ischemic stroke development (among the debatable causes of this outcome may be aspirin resistance). Aspirin intake has a more effective antiaggregate influence on women than men: the intensity of platelet aggregation in women plasma was lower using all agonists, while in men, on the contrary, platelets responded by reaching levels above normal. 4. The patients, who experienced ischemic stroke... [to full text]

In the absence of ligand, certain growth factor receptors can be activated via G protein-coupled receptor (GPCR) activation in a process termed transactivation. Serotonin (5-HT) receptors can transactivate the receptor tyrosine kinase (RTK) platelet-derived growth factor (PDGF) ?? receptors in smooth muscle cells, but it is not known if similar pathways occur in neuronal cells. Here, it is shown that 5-HT can transiently increase the phosphorylation of PDGF?? receptors in a time- and concentration-dependent manner in SH-SY5Y neuroblastoma cells. This transactivation pathway was pertussis-toxin sensitive, and was dependent on phospholipase C activity, intracellular calcium signaling and subsequent protein kinase C activation. Exogenous application of non-lethal concentrations of H2O2 induced the phosphorylation of PDGF?? receptors in a concentration-dependent fashion, similar to that observed with 5-HT. Further investigation revealed reactive oxygen species (ROS) production as a necessary component in the transactivation pathway, as scavenging ROS eliminated PDGF?? receptor phosphorylation. NADPH oxidase was determined to be the likely source of ROS given that the NADPH oxidase inhibitors diphenyleneiodonium chloride and apocynin abrogated PDGF?? receptor transactivation. The role of Src tyrosine kinase was also investigated, and its location in this signaling cascade was determined to be downstream of calcium signaling, but upstream of NADPH oxidase activity. In addition, the activation of ERK1/2 in this system was elucidated to be independent of PDGF?? receptor transactivation. Interestingly, 5-HT also transactivated TrkB receptors, another RTK whose function is implicated in clinical depression. Expectedly, the enzymes in this mechanism were consistent with those revealed in 5-HT-to-PDGF?? receptor signaling. This cross-talk between 5-HT and RTKs such as TrkB and PDGF?? receptors identifies a potentially important signaling link between the serotonergic system and neurotrophic factor signaling in neurons that could have implications in mental health disorders including depression. Furthermore, although transactivation pathways are commonly initiated by a GPCR, recent reports have demonstrated that selective serotonin reuptake inhibitors (SSRIs) were able to block 5-HT-induced transactivation of PDGF?? receptors, suggesting that in addition to GPCRs, monoamine transporters may also be involved in RTK transactivation. SH-SY5Y cells pretreated with the SSRI fluoxetine blocked 5-HT-induced transactivation of the PDGF?? receptors, but not PDGF-induced PDGF?? receptor activation. Upon further examination, it was discovered that during the pretreatment period, fluoxetine itself was transiently transactivating the PDGF?? receptor via 5-HT2 receptors. By the end of the pretreatment period, the effects of fluoxetine on PDGF?? receptor phosphorylation had returned to baseline, and a subsequent transactivating stimulus (5-HT) failed to ???re-transactivate??? the PDGF?? receptor. Additional investigations demonstrated that 5-HT pretreatment can block dopamine-induced PDGF?? receptor transactivation, but not PDGF-induced PDGF?? receptor activation. This is the first demonstration of the heterologous desensitization of an RTK via a transactivation pathway, and this phenomenon is specific for transactivation pathways because in all cases the PDGF?? receptor ligand PDGF-BB was able to directly stimulate receptor activity in spite of GPCR agonist pretreatment. Heterologous desensitization in transactivation signaling reveals a previously unknown short-term ???blackout??? period wherein no further transactivation signaling can occur to potentially exploit the mitogenic effects of RTK activation.

Thrombosis formation upon rupture or erosion of an atherosclerotic plaque can lead to occlusion of arteries. An occlusive thrombus is the most common cause of clinical events such as angina, myocardial infarction, ischemic attacks and strokes. Occlusive thrombi can cause ischemic cardiac arrest in less than an hour. Thrombosis formation requires rapid platelet accumulation rates exceeding thrombosis lysis and embolization rates. Hemodynamics greatly affects platelet accumulation rate through affecting platelet transport to the surface of a growing thrombus. The presence of red blood cells (RBCs) in blood increases platelet transport rate by several orders of magnitude compared to transport due to Brownian motion. Margination of platelets towards the vessel walls also results in higher platelet concentration at the RBC-depleted layer relative to the bulk. In this thesis, we studied the effects of hemodynamics on thrombus growth. We investigated the effects of important flow and particle properties on margination of particles in RBC suspensions by direct numerical simulation (DNS) of cellar blood flow. We derived a scaling law for margination length. Based on this scaling law, margination length increases cubically with channel height and is independent of shear rate. Using DNS, we verified the proposed scaling law for margination length in straight channels. We also showed that rigidity and size both lead to particle margination. We show that platelet margination can be explained by RBC-enhanced shear-induced diffusion of platelets in the RBC-filled region combined with platelet trapping in the RBC-free region. A simple continuum model is introduced based on the proposed mechanism. Using an experimental correlation for effective diffusivity in blood, the continuum model can recover experimental results from the literature over a wide range of tube diameters. We created an in vitro experimental model of thrombosis with and without RBCs. Surprisingly, we found that rapid thrombus growth does not require enhanced platelet transport in the presence of RBCs at high shear. Instead, our results suggest that thrombus growth rate at high shear is dependent on the availability of vWF-A1 domains as opposed to convective transport of platelets. Finally, we obtained empirical correlations for thrombus growth and lag time based on flow parameters by using an in vitro model of thrombosis. We developed a simple model for predicting thrombus formation using the obtained empirical correlations. We demonstrated the capability of the model in predicting thrombus formation over a wide range of experimental geometries. This model may be useful for designing blood-contacting devices to avoid unwanted thrombosis.

We have recently established that aberrant alkylacylglycerophosphocholine metabolism results in the increased tissue concentration of platelet activating factors (PAFs) in the temporal cortex of Alzheimer Disease (AD) patients and in TgCRND8 mice over-expressing mutant human amyloid precursor protein. PAF lipids activate a G-protein coupled receptor (PAFR) reported to be expressed by microglia and subsets of neurons in rat. It is not known whether this same expression pattern is recapitulated in mice however, as the expression has only been inferred by use of pharmacological PAFR antagonists, many of which impact on both PAFR-dependent and PAFR-independent signalling pathways. PAFR plays a role in long term potentiation (LTP) induction in rats. PAFR has also been implicated in behavioural indices of spatial learning and memory in rats. Contradictory reports using mice provide ambiguity regarding the role of PAFR in LTP induction in mice. To assess whether PAFR is expressed in murine neurons, I localized PAFR mRNA in wild-type C57BL/6 mice using PAFR KO mice as a negative control. I further showed that the loss of PAFR did not impair learning and memory although this assessment must be considered preliminary as the behavioural test employed was not optimized to detect changes in learning and memory of C57BL/6 mice over time adequately.Finally, I showed that the loss of PAFR in TgCRND8 mouse model of AD had no impact upon Aβ plaque number. My observations suggest that PAFR is restricted to microglial-like cells in mouse hippocampus and as such, it may not play a role in learning and memory.

Aβ oligomer-induced neurotoxicity has become an important area of therapeutic development in treating Alzheimer’s disease. Platelet-derived growth factor (PDGF) has been shown to be able to protect neurons against several neuronal insults such as ischemia and HIV1 toxin induced cytotoxicity. These neuroprotective effects correlate well with our previous results that demonstrate the neuroprotective effects of PDGF-BB, one of the PDGF receptor ligand subtypes, against NR2B containing NMDA receptor induced excitotoxicity, a possible underlying cause of Aβ oligomer induced synaptic dysfunction and neuronal death. This project examines the neuroprotective effect of PDGF-BB against Aβ1-42 oligomer induced cytotoxicity in both SH-SY5Y cells and primary hippocampal neurons. Cell viability was monitored by MTT assay and the affected signaling pathways were examined using pharmacological methods and Western blotting. The results demonstrated that Aβ1-42 oligomer elicited a dose-dependent toxicity with a sign of saturation at higher dosages, PDGF-BB failed to protect neurons against Aβ1-42 oligomer induced cytotoxicity. In contrast, Aβ1-42 oligomers strongly inhibit PDGF-BB induced mitogenesis in both SH-SY5Y cells and primary neurons. Further investigation using Western blotting to measure PDGF receptor expression and phosphorylation in SH-SY5Y cells showed that Aβ1-42 oligomer can inhibit PDGF-BB induced phosphorylation of PDGF β-receptor on Tyr1021, a site that is crucial for PLCγ mediated mitogenesis. These findings not only explained the poor neuroprotective effect elicited by PDGF-BB against Aβ1-42 oligomers, but also led to a novel hypothesis that Aβ1-42 oligomer may interfere with neurotrophic factor induced neuronal survival, either selectively or perhaps globally. Further exploration on this hypothesis will be able to shed light on this potentially novel mechanism of pathogenesis in Alzheimer’s disease.