Immunospecific analysis of in vitro and ex vivo surface-immobilized protein complex.

Biomaterials used for blood contacting devices are inherently thrombogenic. Antithrombotic agents can be used as surface modifiers on biomaterials to reduce thrombus formation on the surface and to maintain device efficacy. For quality control and to assess the effectiveness of immobilization strategies, it is necessary to quantify the surface-immobilized antithrombotic agent directly. There are limited methods that allow direct quantification on device surfaces such as catheters. In this study, an enzyme immunoassay (EIA) has been developed to measure the density of a synthetic antithrombin-heparin (ATH) covalent complex immobilized on a catheter surface. The distribution of the immobilized ATH was further characterized by an immunohistochemical assay. This analyte-specific EIA is relatively simple and has high throughput, thus providing a tool for quantitative analysis of biomaterial surface modifications. These methods may be further modified to evaluate plasma proteins adsorbed and immobilized on various biomaterial surfaces of complex shapes, with a range of bioactive functionalities, as well as to assess conformational changes of proteins using specific antibodies.

Protocol for the MATCH study (Mindfulness and Tai Chi for cancer health): A preference-based multi-site randomized comparative effectiveness trial (CET) of Mindfulness-Based Cancer Recovery (MBCR) vs. Tai Chi/Qigong (TCQ) for cancer survivors.

PURPOSE: A growing number of cancer survivors suffer high levels of distress, depression and stress, as well as sleep disturbance, pain and fatigue. Two different mind-body interventions helpful for treating these problems are Mindfulness-Based Cancer Recovery (MBCR) and Tai Chi/Qigong (TCQ). However, while both interventions show efficacy compared to usual care, they have never been evaluated in the same study or directly compared. This study will be the first to incorporate innovative design features including patient choice while evaluating two interventions to treat distressed cancer survivors. It will also allow for secondary analyses of which program best targets specific symptoms in particular groups of survivors, based on preferences and baseline characteristics. METHODS AND SIGNIFICANCE: The design is a preference-based multi-site randomized comparative effectiveness trial. Participants (N=600) with a preference for either MBCR or TCQ will receive their preferred intervention; while those without a preference will be randomized into either intervention. Further, within the preference and non-preference groups, participants will be randomized into immediate intervention or wait-list control. Total mood disturbance on the Profile of mood states (POMS) post-intervention is the primary outcome. Other measures taken pre- and post-intervention and at 6-month follow-up include quality of life, psychological functioning, cancer-related symptoms and physical functioning. Exploratory analyses investigate biomarkers (cortisol, cytokines, blood pressure/Heart Rate Variability, telomere length, gene expression), which may uncover potentially important effects on key biological regulatory and antineoplastic functions. Health economic measures will determine potential savings to the health system.

Comparison of recombinant and plasma-derived antithrombin biodistribution in a rabbit model.

Antithrombin (AT) is a native plasma protein that acts as the main inhibitor of enzymes generated by the coagulation cascade. In extreme thrombotic conditions, consumption of plasma AT can make treatment with AT-associated heparin therapies less effective. Supplementation with recombinant human AT (rhAT) has shown promise but altered pharmacokinetics were observed when comparing stable heparin complexes of the plasma-derived AT (pAT) and rhAT proteins. To understand the differential clearance mechanisms, biodistribution of rhAT and pAT was determined. (125)I-labelled ATryn (rhAT) or Kybernin P (pAT) was intravenously injected into rabbits. At various time points, animals were sacrificed and organs analysed for bound radioactivity. (131)I-albumin, injected shortly before termination, was used as a marker for trapped blood. Levels of circulating protein + metabolites were significantly less for rhAT than pAT (p < 0.001) and removal of acid soluble fragments confirmed that differences were due to more rapid rhAT clearance. More rhAT (28% dose) than pAT (3% dose) was liver-associated by the earliest time points, corresponding to elevated rhAT degradation products in urine/feces. However, at intermediate times (4 hours), rhAT showed significantly increased arterial and venous uptake over pAT (p

Binding of heparin to plasma proteins and endothelial surfaces is inhibited by covalent linkage to antithrombin.

Unfractionated heparin (UFH) and low molecular weight heparin (LMWH) are used for prophylaxis and treatment of thrombosis. However, UFH has a short plasma half-life and variable anticoagulant response in vivo due to plasma or vessel wall protein binding and LMWH has a decreased ability to inactivate thrombin, the pivotal enzyme in the coagulation cascade. Covalent linkage of antithrombin to heparin gave a complex (ATH) with superior anticoagulant activity compared to UFH and LMWH, and longer intravenous half-life compared to UFH. We found that plasma proteins bound more to UFH than ATH, and least to LMWH. Also, UFH bound significantly more to endothelial cells than ATH, with 100% of UFH and 94% of ATH binding being on the cell surface and the remainder was endocytosed. Competition studies with UFH confirmed that ATH binding was likely through its heparin moiety. These findings suggest that differences in plasma protein and endothelial cell binding may be due to available heparin chain length. Although ATH is polydisperse, the covalently-linked antithrombin may shield a portion of the heparin chain from association with plasma or endothelial cell surface proteins. This model is consistent with ATH's better bioavailability and more predictable dose response.