Antidote-Controlled Platelet Inhibition Targeting von Willebrand Factor with Aptamers

Thrombus formation is initiated by platelets and leads to cardiovascular, cerebrovascular, and peripheral vascular disease, the leading causes of morbidity and mortality in the Western world. A number of antiplatelet drugs have improved clinical outcomes for thrombosis patients. However, their expanded use, especially in surgery, is limited by hemorrhage. Here, we describe an antiplatelet agent that can have its activity controlled by a matched antidote. We demonstrate that an RNA aptamer targeting von Willebrand factor (VWF) can potently inhibit VWF-mediated platelet adhesion and aggregation. By targeting this important adhesion step, we show that the aptamer molecule can inhibit platelet aggregation in PFA-100 and ristocetin-induced platelet aggregation assays. Furthermore, we show that a rationally designed antidote molecule can reverse the effects of the aptamer molecule, restoring platelet function quickly and effectively over a clinically relevant period. This aptamer-antidote pair represents a reversible antiplatelet agent inhibiting a platelet specific pathway. Furthermore, it is an important step towards creating safer drugs in clinics through the utilization of an antidote molecule.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/63204/1/oli.2007.0089.pd

Aptamers as molecular recognition elements for electrical nanobiosensors

Recent advances in nanotechnology have enabled the development of nanoscale sensors that outperform conventional biosensors. This review summarizes the nanoscale biosensors that use aptamers as molecular recognition elements. The advantages of aptamers over antibodies as sensors are highlighted. These advantages are especially apparent with electrical sensors such as electrochemical sensors or those using field-effect transistors

Intra-Accumbens Injection of a Dopamine Aptamer Abates MK-801-Induced Cognitive Dysfunction in a Model of Schizophrenia

Systemic administration of the noncompetitive NMDA-receptor antagonist, MK-801, has been proposed to model cognitive deficits similar to those seen in patients with schizophrenia. The present work investigated the ability of a dopamine-binding DNA aptamer to regulate these MK-801-induced cognitive deficits when injected into the nucleus accumbens. Rats were trained to bar press for chocolate pellet rewards then randomly assigned to receive an intra-accumbens injection of a DNA aptamer (200 nM; n = 7), tris buffer (n = 6) or a randomized DNA oligonucleotide (n = 7). Animals were then treated systemically with MK-801 (0.1 mg/kg) and tested for their ability to extinguish their bar pressing response. Two control groups were also included that did not receive MK-801. Data revealed that injection of Tris buffer or the random oligonucleotide sequence into the nucleus accumbens prior to treatment with MK-801 did not reduce the MK-801-induced extinction deficit. Animals continued to press at a high rate over the entire course of the extinction session. Injection of the dopamine aptamer reversed this MK-801-induced elevation in lever pressing to levels as seen in rats not treated with MK-801. Tests for activity showed that the aptamer did not impair locomotor activity. Results demonstrate the in vivo utility of DNA aptamers as tools to investigate neurobiological processes in preclinical animal models of mental health disease

A Grafting Strategy for the Design of Improved G-Quadruplex Aptamers and High-Activity DNAzymes

Nucleic acid aptamers are generally obtained by in vitro selection. Some have G-rich consensus sequences with ability to fold into the four-stranded structures known as G-quadruplexes. A few G-quadruplex aptamers have proven to bind hemin to form a new class of DNAzyme with the peroxidase-like activity, which can be significantly promoted by appending an appropriate base-pairing duplex onto the G-quadruplex structures of aptamers. Knowing the structural role of base pairing, here we introduce a novel grafting strategy for the design of improved G-quadruplex aptamers and high-activity DNAzymes. To demonstrate this strategy, three existing G-quadruplex aptamers are chosen as the first generation. A base-pairing DNA duplex is grafted onto the G-quadruplex motif of the first generation aptamers. Consequently, three new aptamers with the quadruplex/duplex DNA structures are produced as the second generation. The hemin-binding affinities and DNAzyme functions of the second generation aptamers are characterized and compared with the first generation. The results indicate three G-quadruplex aptamers obtained by the grafting strategy have more excellent properties than the corresponding original aptamers. Our findings suggest that, if the structures and functions of existing aptamers are thoroughly known, the grafting strategy can be facilely utilized to improve the aptamer properties and thereby producing better next-generation aptamers. This provides a simple but effective approach to the design of nucleic acid aptamers and DNAzymes

Iron Behaving Badly: Inappropriate Iron Chelation as a Major Contributor to the Aetiology of Vascular and Other Progressive Inflammatory and Degenerative Diseases

The production of peroxide and superoxide is an inevitable consequence of aerobic metabolism, and while these particular "reactive oxygen species" (ROSs) can exhibit a number of biological effects, they are not of themselves excessively reactive and thus they are not especially damaging at physiological concentrations. However, their reactions with poorly liganded iron species can lead to the catalytic production of the very reactive and dangerous hydroxyl radical, which is exceptionally damaging, and a major cause of chronic inflammation. We review the considerable and wide-ranging evidence for the involvement of this combination of (su)peroxide and poorly liganded iron in a large number of physiological and indeed pathological processes and inflammatory disorders, especially those involving the progressive degradation of cellular and organismal performance. These diseases share a great many similarities and thus might be considered to have a common cause (i.e. iron-catalysed free radical and especially hydroxyl radical generation). The studies reviewed include those focused on a series of cardiovascular, metabolic and neurological diseases, where iron can be found at the sites of plaques and lesions, as well as studies showing the significance of iron to aging and longevity. The effective chelation of iron by natural or synthetic ligands is thus of major physiological (and potentially therapeutic) importance. As systems properties, we need to recognise that physiological observables have multiple molecular causes, and studying them in isolation leads to inconsistent patterns of apparent causality when it is the simultaneous combination of multiple factors that is responsible. This explains, for instance, the decidedly mixed effects of antioxidants that have been observed, etc...Comment: 159 pages, including 9 Figs and 2184 reference

The interstitium in cardiac repair: role of the immune-stromal cell interplay

Cardiac regeneration, that is, restoration of the original structure and function in a damaged heart, differs from tissue repair, in which collagen deposition and scar formation often lead to functional impairment. In both scenarios, the early-onset inflammatory response is essential to clear damaged cardiac cells and initiate organ repair, but the quality and extent of the immune response vary. Immune cells embedded in the damaged heart tissue sense and modulate inflammation through a dynamic interplay with stromal cells in the cardiac interstitium, which either leads to recapitulation of cardiac morphology by rebuilding functional scaffolds to support muscle regrowth in regenerative organisms or fails to resolve the inflammatory response and produces fibrotic scar tissue in adult mammals. Current investigation into the mechanistic basis of homeostasis and restoration of cardiac function has increasingly shifted focus away from stem cell-mediated cardiac repair towards a dynamic interplay of cells composing the less-studied interstitial compartment of the heart, offering unexpected insights into the immunoregulatory functions of cardiac interstitial components and the complex network of cell interactions that must be considered for clinical intervention in heart diseases

A Novel Case of Transverse Sinus Stenting and Ticagrelor Use During Pregnancy for Idiopathic Intracranial Hypertension

Background Idiopathic intracranial hypertension (IIH) is prevalent among the US population, with exacerbation of symptoms during pregnancy. Transverse sinus stenting is a new effective treatment for IIH. Stenting is avoided in pregnancy largely due to the requirement of dual antiplatelet therapy. Methods We present a case of a pregnant patient in her first trimester with signs and symptoms of fulminant IIH, including progressive visual loss, who underwent placement of a transverse sinus stent and administration of dual antiplatelet therapy with ticagrelor. Results Ticagrelor was administered pre–operatively. The patient underwent venous sinus stenting for fulminant IIH. There were no complications. She had resolution of symptoms and underwent cesarean delivery without issues. Her child displayed no signs and symptoms of abnormalities. Conclusion Venous sinus stenting for IIH can be considered in pregnant patients presenting with new or worsening IIH with associated papilledema. The use of ticagrelor did not lead to any adverse outcomes for the patient or the fetus in our case

A canine thromboembolic model of anterior circulation large vessel occlusion stroke

Purpose: To develop a large animal preclinical model of thromboembolic stroke with stable, protracted large vessel occlusion (LVO) utilizing an autologous clot. Materials and methods: A reproducible canine model of large vessel occlusion stroke was established by endovascular placement of an autologous clot into the middle cerebral artery (MCA) of six adult hounds and confirmed using digital subtraction angiography (DSA). Infarct volume and evidence of hemorrhage were determined by magnetic resonance imaging (MRI) 7 h after occlusion and Thrombolysis in Cerebral Infarction scale (TICI) was assessed before and after clot placement and at 1, 6, 7, and 9 h after middle cerebral artery occlusion (MCAO). Heart rate (HR) and blood pressure (BP) were monitored continuously and invasively through an arterial sheath throughout the procedures and complete blood count and blood gas analysis completed at time of sacrifice. Histopathological findings at time of sacrifice were used to confirm stroke volume and hemorrhage. Results: MCAO with resulting TICI 0 flow was observed in all six animals, verified by serial DSA, and lack of collateral flow persisted for 9 h after clot placement until time of sacrifice. The mean infarct volume was 47.0 ± 6.7% of the ipsilateral hemisphere and no events of spontaneous recanalization or clot autolysis were observed. Conclusion: We demonstrate a thromboembolic canine model of MCAO that is both feasible and results in consistent infarct volumes to generate a clinically relevant LVO. This model is important to evaluate treatment of LVO in acute ischemic stroke (AIS) outside the established 4.5 h recombinant tissue plasminogen activator (rTPA) therapeutic window utilizing a prolonged occlusive thrombus