Feasibility of opportunistic colorectal cancer screening of hospitalized patients in tertiary care and community hospitals.

BACKGROUND: Population screening for colorectal cancer (CRC) remains low, requiring alternative approaches for increasing participation. Opportunistic screening of hospitalized patients may aid in increasing uptake rates. OBJECTIVE: To assess whether inpatients can be recruited for opportunistic CRC screening using fecal immunochemical testing (FIT). METHODS: Inpatient charts were prospectively reviewed for study eligibility on admission of patients to the medical wards of 3 hospitals in Winnipeg, Canada. Eligible patients were approached for participation and consent. Inoculated FIT specimen collection tubes were sent to the hospital laboratory for testing. Patients with positive FIT results received a follow-up colonoscopy. RESULTS: In total, 1542 inpatient charts were screened for eligibility; 53 patients were identified for enrollment (51.9% were male; median age, 59 years), of whom 13 patients consented to participate but only 7 provided a stool specimen. One of those 7 patients had a positive FIT result. The overall screening rate was low, at 0.45%. The primary reason for exclusion of patients was age (outside of the range of 50-75 years), followed by patients having recent gastrointestinal bleeding and/or known intestinal diseases. CONCLUSIONS: Our data suggest that it is infeasible to recruit inpatients for opportunistic CRC screening in routine clinical practice.

Every Tube Counts: reducing extra tubes drawn in the emergency department.

A common practice exists in hospitals where extra tubes of blood are collected for possible add-on testing, this practice contributes to wastage of consumables. Baseline estimates from a 5-month local lab information system audit revealed that ~65 extra tubes per day were being collected, with an additional 2-week manual audit of all extra tubes received in the laboratory confirming the practice. The audits showed that the majority of the tubes (~99%) were being drawn from the adult emergency department (ED). Furthermore, only 5% of the extra tubes were being used for add-on testing, whereas the remaining tubes had no testing performed on them and were discarded at the end of the day. This translates to over 23 000 extra tubes being wasted annually.After initial discussion with ED leadership, the practice was identified as primarily nurse driven. An educational intervention was created and entitled 'Every Tube Counts', with the aim to reduce extra tube collections in the adult ED by 50% within the first month of intervention. First, a memo with initial findings and a request to stop the practice of extra tube collection was sent out to all ED staff. After 2 weeks of additional data collection, it was noticed that extra tubes were still being collected. A second intervention, which consisted of another communication and utilisation of nurse educators to disseminate the information to nursing staff, saw a remarkable ~80% reduction in collection of extra tubes in the following few months after the second intervention. The practice was followed for an additional 15 months, which saw a slight increase of extra tube collections over time with a levelling off towards the latter period of the study. However, the target goal was maintained over the entire study period.

Long term stability of lactate in uncentrifuged sodium fluoride/potassium oxalate blood collection tubes.

BACKGROUND: Delayed time from collection to centrifugation may cause erroneously high lactate levels in vitro (from continued blood cell metabolism under anaerobic conditions in the collection tube) if not collected in appropriate collection devices, consequently increasing the risk for inappropriate patient care or harm. We undertook a study to determine the turnaround time for lactate testing in a tertiary care setting and also performed short- and long-term lactate stability studies in blood collected in sodium fluoride/potassium oxalate (NaF/KOx) collection tubes. METHODS: The hospital lab information system was mined for 6 months to determine patient samples that may have exceeded the time from collection-to-receival in lab of 15-min. Lactate stability was evaluated in unspun NaF/KOx collection tubes at 15 min intervals for to 2 h; and separately at 2, 6, 12, 24, and 48-h post-collection. RESULTS: A total of 8,929 plasma samples were collected in 6 months, and 1/3 were not received in the lab within 15 min from collection. In NaF/KOx additive, lactate levels had minor increases over 2 h, and incremental increases at an average rate of 0.0035 mmol/L/h over 48 h with maximum increase of 9.8% at 48 h. However, the average change across all time points were within local allowable performance goals (at ≤4 mmol/L ± 0.5 mmol/L; at >4 mmol/L ± 12%). CONCLUSION: A small proportion of lactate specimens may experience delay in processing. Although lactate levels may incrementally increase over 48-h at room temperature in unspun NaF/KOx collection tubes, the changes may not be clinically impactful.

Lack of interference of metronidazole with Roche cobas c502 and c702 chemistry tests.

OBJECTIVE: The antibiotic metronidazole has been suggested to absorb light at a wavelength range commonly used in spectrophotometric assays. We sought to determine if any of the spectrophotometric assays used in our core laboratory would be susceptible to clinically significant interference from metronidazole in blood-based patient specimens. METHODS: Following characterization of the absorbance spectrum for metronidazole, spectrophotometric assays involving either main or subtraction wavelengths that might be susceptible to interference from metronidazole were identified. A total of 24 chemistry tests performed on Roche cobas c502 and/or c702 instruments were evaluated for interference from metronidazole. For each assay, two pools of leftover patient serum, plasma, or whole blood specimens containing the analyte of interest at clinically relevant concentrations were prepared. Each pool was spiked with metronidazole at a final concentration of 200 mg/L (1169 µmol/L) or 10 mg/L (58 µmol/L) or the same volume of water as a control, with triplicate samples for each group. The difference in the measured analyte concentration between experimental and control groups was then compared against the total allowable error for each assay to determine whether clinically significant interference had occurred. RESULTS: There was no significant interference observed with Roche chemistry tests due to the presence of metronidazole. CONCLUSION: This study provides reassurance that metronidazole is not interfering with the chemistry assays used in our core laboratory. Interference from metronidazole may be a historical problem and current spectrophotometric assays may not be susceptible due to improvements in assay design.

Co-administration of FVIII with IVIG reduces immune response to FVIII in hemophilia A mice.

Hemophilia A is an X-linked recessive congenital bleeding disorder. Exogenous infusion of FVIII is the treatment of choice, and the development of immunoglobulins against FVIII (inhibitors) remains the major challenge in clinical management of the disease. Here, we investigated the effect of co-administration of FVIII with intravenous immunoglobulin (IVIG) on the development of inhibitors in previously untreated hemophilia A mice. A group of hemophilia A mice (C57BL/6FVIII-/-) received weekly injections of recombinant human FVIII (rFVIII) for twelve consecutive weeks while a second group received co-injections of rFVIII + IVIG. An in-house enzyme-linked immunosorbent assay (ELISA) was designed to detect antibodies to rFVIII. Every mouse in the first group developed antibodies to rFVIII. In contrast, mice treated with rFVIII + IVIG showed significantly lower antibody titers. Interestingly, when co-administration of IVIG was discontinued after 12 weeks in some mice (rFVIII continued), these mice experienced an increase in antibody titer. In contrast, mice that continued to receive rFVIII + IVIG retained significantly lower titers. In conclusion, prophylactic rFVIII co-administration with IVIG modulated the immune response to FVIII and resulted in decreased anti-FVIII antibody titer. These findings suggest that co-injection therapy with IVIG could potentially be effective in the management of hemophilia A patients at risk of inhibitor development.

Effect of unfractionated heparin and low molecular weight heparin on the clotting of platelet-reduced whole blood: an in-vitro study utilizing thromboelastography.

Treatment of venous thromboembolism with concomitant thrombocytopenia is challenging. The platelet threshold for safe administration of anticoagulants is under debate, with minimum platelet count of 50 × 109/l being recommended as the safe cutoff. However, some evidence suggests administration of anticoagulants may still be safe at platelet levels of 30 × 109/l. Therefore, we developed an in-vitro thromboelastography (TEG) study to examine the effect of therapeutic or prophylactic levels of unfractionated heparin (UFH) and low molecular weight heparin (LMWH) on the clotting profile of platelet-reduced whole blood. Using magnetic bead-based antibody chromatography, platelets were removed to achieve platelet-depleted blood (<10 × 109/l of platelets). Platelet-depleted blood was then mixed with whole blood to produce blood samples with platelet counts of 30 × 109, 50 × 109 and 150 × 109/l. These blood samples were incubated with therapeutic or prophylactic levels of UFH or LMWH in disposable TEG cups. Clotting was initiated with 10 mmol/l calcium and optimized tissue factor levels for each anticoagulant used (2.25 pmol/l for UFH and 2.05 pmol/l for LMWH). Clotting was monitored by TEG at 37 °C for 180 min. The following TEG parameters were evaluated: R (time to clot), maximum amplitude (strength of clot) and area under the curve in 15 min (overall speed and strength of the clot at 15 min of clotting). No statistically significant differences were observed between platelet counts of 30 × 109 and 50 × 109/l for R, maximum amplitude or area under the curve in 15 min for most of the therapeutic and prophylactic doses of UFH and LMWH tested in this study. Use of anticoagulants compromised all of the TEG parameters relative to a normal platelet count of 150 × 109/l, in a dose dependent manner. The current study demonstrates that in-vitro clotting is impaired with use and increasing doses of anticoagulants. Despite this observation, we did not observe a significant difference in clotting between platelet levels of 30 × 109 and 50 × 109/l. Overall, this work provides further insight in the debated use of anticoagulants in patients with venous thromboembolism and concomitant thrombocytopenia, and provides support for possible use of anticoagulants at lower platelet thresholds.

Biochemistry tests in hospitalized COVID-19 patients: Experience from a Canadian tertiary care centre.

BACKGROUND: Coronavirus Disease 2019 (COVID-19) has variable clinical presentation, from asymptomatic to severe disease leading to death. Biochemical markers may help with management and prognostication of COVID-19 patients; however, their utility is still under investigation. METHODS: A retrospective study was conducted to evaluate alanine aminotransferase, C-reactive protein (CRP), ferritin, lactate, and high sensitivity troponin T (TnT) levels in 67 patients who were admitted to a Canadian tertiary care centre for management of COVID-19. Logistic, cause-specific Cox proportional-hazards, and accelerated failure time regression modelling were performed to assess the associations of initial analyte concentrations with in-hospital death and length of stay in hospital; joint modelling was performed to assess the associations of the concentrations over the course of the hospital stay with in-hospital death. RESULTS: Initial TnT and CRP concentrations were associated with length of stay in hospital. Eighteen patients died (27%), and the median initial TnT concentration was higher in patients who died (55 ng/L) than those who lived (16 ng/L; P < 0.0001). There were no survivors with an initial TnT concentration > 64 ng/L. While the initial TnT concentration was predictive of death, later measurements were not. Only CRP had prognostic value with both the initial and subsequent measurements: a 20% increase in the initial CRP concentration was associated with a 14% (95% confidence interval (CI): 1-29%) increase in the odds of death, and the hazard of death increased 14% (95% CI: 5-25%) for each 20% increase in the current CRP value. While the initial lactate concentration was not predictive of death, subsequent measurements were. CONCLUSION: CRP, lactate and TnT were associated with poorer outcomes and appear to be useful biochemical markers for monitoring COVID-19 patients.

Rivaroxaban and dabigatran did not affect clotting profiles in plasma reconstituted with varying levels of autologous platelets to the same degree as heparin when evaluated using thromboelastography.

OBJECTIVE: Current recommendations for treating patients with thromboembolism and concomitant thrombocytopenia are based on anecdotal data and expert opinion, rather than clinical studies. Our aim was to use an in-vitro model employing thromboelastography (TEG) to evaluate clot formation as a surrogate indicator of clinical tendency to hemorrhage, and investigate the interactions of plasma at varying concentrations of platelets in the presence of anticoagulants. METHODS: Platelet-rich and platelet-poor plasma isolated from whole blood were mixed together to obtain platelet concentrations ranging from less than 10-150 × 10 platelets/l. Clotting was initiated with tissue factor and measured by TEG. RESULTS: Different tissue factor concentrations were required to model clinical clotting profiles for plasma that contained heparin (UFH), low molecular weight heparin (LMWH), or fondaparinux. No tissue factor was required for rivaroxaban or dabigatran-clotting reactions. The time to initiate coagulation (R) was significantly delayed at platelet concentrations less than 30 × 10/l for UFH and LMWH, less than 20 × 10/l for fondaparinux, and less than 10 × 10/l for rivaroxaban and dabigatran. The strength of the clot was significantly compromised at all platelet concentrations in the presence of UFH, LMWH or fondaparinux. In contrast, rivaroxaban and dabigatran compromised clot strength at platelet concentrations less than 10 × 10/l. CONCLUSION: All anticoagulants tested compromised coagulation at specific platelet concentration thresholds. Rivaroxaban and dabigatran had reduced impact on clot formation at low-platelet concentrations compared with heparinoids, suggesting that the factor-specific inhibitors may be more favorable than traditional heparin-based treatment of thromboembolism in the presence of thrombocytopenia.

Transient early increase in thyroglobulin levels post-radioiodine ablation in patients with differentiated thyroid cancer.

OBJECTIVES: Treatment of differentiated thyroid cancer (DTC) includes surgical thyroidectomy and, in most cases, radioactive iodine (RAI) ablation. Measurement of serum thyroglobulin (Tg) levels is used for assessing disease burden and identifying persistent-recurrent DTC. This prospective study determined the Tg profile before and after RAI-ablation in patients with DTC. DESIGN AND METHODS: Fifty-five DTC patients with complete resection received RAI-ablation and were assessed for Tg at baseline (non-stimulated), pre-ablation (stimulated), 7 days post-ablation (stimulated) and at 6 months (stimulated). Stimulation of Tg was achieved by thyroid hormone withdrawal to achieve serum thyroid stimulating hormone (TSH) ≥30 mU/L. Thyroid remnant size was estimated from whole body scintigraphy. Similar protocols were implemented for nine patients with incomplete resection/metastatic disease for comparison. RESULTS: Mean stimulated Tg levels for DTC patients with complete resection at 7 days post-RAI increased 13-fold from 13.7 to 175.5 µg/L (p<0.0001), and the Tg levels reduced to 2.3 µg/L (p<0.0001 versus post-RAI) by follow-up. None of the patients had recurrence of disease. For the nine patients with incomplete resection/metastases, Tg levels were higher throughout compared to the patients with complete resection. There was no increase in Tg between pre- and post-RAI. We did not observe a significant correlation between the remnant size and Tg increase. CONCLUSIONS: This study confirms a prominent transient early increase in Tg post-RAI ablation in DTC patients with complete resection, with the Tg levels falling below baseline by 6 months. This is presumed to reflect RAI-induced thyroid tissue destruction/inflammation with subsequent release of Tg from the thyroid remnant. Recognizing this transient phenomenon is important for post-ablation Tg interpretation and patient management.

Phenotype presentation for a novel mutation affecting a conserved cysteine residue in exon 63 of fibrillin-1 (Cys2633Arg).

Marfan syndrome is an autosomal dominant disease caused by mutations in the gene encoding for fibrillin-1 (FBN1). More than 1,000 FBN1 mutations have been identified, which may lead to multiple organ involvement, particularly of the ocular, skeletal, and cardiovascular systems. Mutations in exons 59-65 have been reported in the past to cause mild Marfan-like fibrillinopathies. We report a family with a mutation in exon 63 that manifests with significant cardiovascular system involvement such as aortic root dilatations, dissection of the aorta, and sudden death at a young age. Genetic analysis revealed that four related individuals are positive for a novel heterozygous Cys2633Arg mutation in exon 63. Their genotype-phenotype profile (based on the revised Ghent nosology) is described. We postulate that the Cys2633Arg mutation may manifest with significant and progressive enlargement of the aortic root, risk of aortic dissections, and minor skeletal abnormalities, without involving the ocular system (i.e., ectopia lentis).

Interactions of heparin and a covalently-linked antithrombin-heparin complex with components of the fibrinolytic system.

Unfractionated heparin (UFH) is used as an adjunct during thrombolytic therapy. However, its use is associated with limitations, such as the inability to inhibit surface bound coagulation factors. We have developed a covalent conjugate of antithrombin (AT) and heparin (ATH) with superior anticoagulant properties compared with UFH. Advantages of ATH include enhanced inhibition of surface-bound coagulation enzymes and the ability to reduce the overall size and mass of clots in vivo. The interactions of UFH or ATH with the components of the fibrinolytic pathway are not well understood. Our study utilised discontinuous second order rate constant (k2) assays to compare the rates of inhibition of free and fibrin-associated plasmin by AT+UFH vs ATH. Additionally, we evaluated the effects of AT+UFH and ATH on plasmin generation in the presence of fibrin. The k2 values for inhibition of plasmin were 5.74 ± 0.28 x 106 M⁻¹ min⁻¹ and 6.39 ± 0.59 x 106 M⁻¹ min⁻¹ for AT+UFH and ATH, respectively. In the presence of fibrin, the k2 values decreased to 1.45 ± 0.10 x 106 M⁻¹ min⁻¹ and 3.07 ± 0.19 x 106 M⁻¹ min⁻¹ for AT+UFH and ATH, respectively. Therefore, protection of plasmin by fibrin was observed for both inhibitors; however, ATH demonstrated superior inhibition of fibrin-associated plasmin. Rates of plasmin generation were also decreased by both inhibitors, with ATH causing the greatest reduction (approx. 38-fold). Nonetheless, rates of plasmin inhibition were 2-3 orders of magnitude lower than for thrombin, and in a plasma-based clot lysis assay ATH significantly inhibited clot formation but had little impact on clot lysis. Cumulatively, these data may indicate that, relative to coagulant enzymes, the fibrinolytic system is spared from inhibition by both AT+UFH and ATH, limiting reduction in fibrinolytic potential during anticoagulant therapy.

Covalently linking heparin to antithrombin enhances prothrombinase inhibition on activated platelets.

Factor (F)Xa within the prothrombinase complex is protected from inhibition by unfractionated heparin (UFH), enoxaparin and fondaparinux. We have developed a covalent antithrombin-heparin complex (ATH) with enhanced anticoagulant activity. We have also demonstrated that ATH is superior at inhibiting coagulation factors when assembled on artificial surfaces. The objective of the present study is to determine the ability of ATH vs AT+UFH to inhibit FXa within the prothrombinase complex when the enzyme complex is assembled on the more native platelet system. Discontinuous inhibition assays were performed to determine final k2-values for inhibition of FXa, FXa within the platelet-prothrombinase, or FXa within prothrombinase devoid of various components. Thrombin generation and plasma clotting was also assayed in the presence of resting/activated platelets ± inhibitors. Protection of FXa was not observed for ATH, whereas a moderate 60% protection was observed for AT+UFH. ATH inhibited platelet-prothrombinase ~4-fold faster than AT+UFH. Relative to intact prothrombinase, ratesfor FXa inhibition by AT+UFH in prothrombinase complexes devoid of either prothrombin (II)/activated platelets/FVa were higher. However, inhibition by AT+UFH of prothrombinase devoid of FII yielded slightly lower rates compared to free FXa inhibition. Thrombin generation and plasma clotting was enhanced with activated platelets, while inhibition was better by ATH compared to AT+UFH, thus suggesting an overall enhanced anticoagulant activity of ATH against platelet-bound prothrombinase complexes.

Inhibition of the prothrombinase complex on red blood cells by heparin and covalent antithrombin-heparin complex.

The role of red blood cells (RBCs) in coagulation is not well understood. Overt exposure of phosphatidylserine on surfaces of RBCs provide docking sites for formation of the prothrombinase complex, which further aids in amplification of coagulation leading to subsequent thrombosis. No studies to date have evaluated heparin inhibition of the RBC-prothrombinase system. Therefore, this study examines the ability of heparin and a covalent antithrombin-heparin complex (ATH) to inhibit the RBC-prothrombinase system. Discontinuous inhibition assays were performed to obtain k2 values for inhibition of free or prothrombinase-bound Xa by antithrombin and unfractionated heparin (AT + UFH) versus ATH. In addition, components of the complex (prothrombin, RBCs or Va) were excluded prior to reaction with inhibitors to investigate potential mechanisms involved. Inhibition of thrombin generation, fibrinogen conversion and plasma clotting by the RBC-prothrombinase system was also examined. Protection of Xa was observed for AT + UFH and not for ATH reactions. Inhibition rates for ATH were significantly faster when compared with AT + UFH results. The greatest impact on Xa inhibition was observed from factor Va omission for both inhibitors. ATH inhibited thrombin generation, fibrinogen conversion and plasma clotting better compared with AT + UFH. This study determined potential control of coagulation contributed by RBCs. Moreover, greater control of coagulation is achieved by covalently linking heparin to AT.

Perilipin family (PLIN) proteins in human skeletal muscle: the effect of sex, obesity, and endurance training.

Proteins that coat the lipid droplets (also known as PAT proteins or perilipin (PLIN) family proteins) have diverse functions that are not well elucidated in many tissues. In skeletal muscle, there is even less known about the functions or characteristics of these proteins or how they might change in response to perturbations that alter both intramyocellular lipid (IMCL) content and fat utilization and oxidation. Therefore, the purpose of this study was to examine the human muscle content and gene expression of the four skeletal muscle PLIN proteins in both lean and obese men and women and how this was changed following a 12-week endurance training protocol. PLIN2-PLIN5 proteins were all more abundant in women than in men (p = 0.037 and p < 0.0001, respectively), consistent with higher IMCL content observed in female skeletal muscle. PLIN5 (previously known as OXPAT) is of particular interest because it has previously been associated primarily with oxidative tissues that rely heavily on fat oxidation for energy production. Although PLIN5 was not different between lean and obese subjects, it was the only PLIN protein to increase in response to endurance training in both sexes. PLIN5 correlated with IMCL volume (p < 0.0001), but in general, the other PLIN proteins did not correlate well with IMCL volume, suggesting that the relationship between lipid accumulation and PLIN family protein content is not a simple one. Although more work is necessary, it is clear that PLIN5 likely plays an important role in IMCL accumulation and oxidation, both of which increase with endurance training in human skeletal muscle.

Mechanism of inhibition of the prothrombinase complex by a covalent antithrombin-heparin complex.

Factor-Xa assembly into the prothrombinase complex decreases its availability for inhibition by antithrombin + unfractionated heparin (AT + UFH). We have developed a novel covalent antithrombin-heparin complex (ATH), with enhanced anticoagulant actions compared with AT + UFH. The present study was performed to extend understanding of the anticoagulant mechanisms of ATH by determining its inhibition of Xa within the critical prothrombinase. Discontinuous inhibition assays were performed to determine final k(2) values for inhibition of Xa. Fluorescent microscopy was conducted to evaluate inhibitor-prothrombinase interactions. The k(2) for inhibition of prothrombinase versus free Xa by AT + UFH was lower, whereas for ATH were much higher. Relative to intact prothrombinase, rates for Xa inhibition by AT + UFH in complexes devoid of prothrombin/vesicles/factor-Va were higher. For ATH, exclusion of prothrombin decreased k(2), removal of vesicles increased k(2) and exclusion of factor-Va gave no effect. While UFH may displace Xa from prothrombinase, Xa is detained within prothrombinase during ATH reactions. We confirm prothrombinase hinders inhibitory action of AT + UFH, whereas ATH is less affected with prothrombin being a key component in the complex responsible for the opposing effects. Overall, the results suggest that covalent linkage between AT-heparin assists access and neutralization of complexed Xa, with concomitant inhibition of prothrombinase function compared with conventional non-conjugated heparin.

Carotid artery dissections: thrombosis of the false lumen.

Carotid artery dissections are the second leading cause of stroke in young adults. The hemostatic response to a dissection involves exposure of the subendothelium to the intravascular environment. Platelet activation/aggregation superimposed by secondary coagulation cascade activity attempts to heal the injury. Failure of the hemostatic response to heal the injury may lead to further rupture of the intimal and medial layers, which allows for the blood to penetrate these layers to create a false lumen. Continued hemorrhaging into the false lumen may result in dissection progression or obstruction of blood supply to the true lumen and downstream blood vessels. The effects of thrombosis in the true versus false lumen may lead to opposite consequences. True lumen clotting may lead to ischemic complications of downstream cerebral vasculature, whereas false lumen clotting may lead to dissection healing. Current information on clinical outcomes and degree of false lumen clotting in a carotid dissection model is limited, and most of the available information on this controversial topic has been inferred from aortic dissections. Therefore in this report we summarize the present state of knowledge of the pathophysiology, detailed hemostatic response to the injury, clinical presentation and treatment of carotid dissections. We also emphasize the need for future studies to investigate the degree of false lumen clotting on the clinical outcomes of carotid dissections.

17ß-estradiol attenuates exercise-induced neutrophil infiltration in men.

17ß-estradiol (E2) attenuates exercise-induced muscle damage and inflammation in some models. Eighteen men completed 150 eccentric contractions after random assignment to placebo (Control group) or E2 supplementation (Experimental group). Muscle biopsies and blood samples were collected at baseline, following 8-day supplementation and 3 h and 48 h after exercise. Blood samples were analyzed for sex hormone concentration, creatine kinase (CK) activity and total antioxidant capacity. The mRNA content of genes involved in lipid and cholesterol homeostasis [forkhead box O1 (FOXO1), caveolin 1, and sterol regulatory element binding protein-2 (SREBP2)] and antioxidant defense (SOD1 and -2) were measured by RT-PCR. Immunohistochemistry was used to quantify muscle neutrophil (myeloperoxidase) and macrophage (CD68) content. Serum E2 concentration increased 2.5-fold with supplementation (P < 0.001), attenuating neutrophil infiltration at 3 h (P < 0.05) and 48 h (P < 0.001), and the induction of SOD1 at 48 h (P = 0.02). Macrophage density at 48 h (P < 0.05) and SOD2 mRNA at 3 h (P = 0.01) increased but were not affected by E2. Serum CK activity was higher at 48 h for both groups (P < 0.05). FOXO1, caveolin 1 and SREBP2 expression were 2.8-fold (P < 0.05), 1.4-fold (P < 0.05), and 1.5-fold (P < 0.001) and higher at 3 h after exercise with no effect of E2. This suggests that E2 attenuates neutrophil infiltration; however, the mechanism does not appear to be lesser oxidative stress or membrane damage and may indicate lesser neutrophil/endothelial interaction.

Binding of heparin to metals.

Heparin is a major prophylactic and treatment agent for thrombosis. Structurally, this anticoagulant is a polydisperse, highly negatively charged polysaccharide mixture that contains a variable density of sulfate group substituents per molecule. Previous study has shown that heparin molecules have a high affinity for a wide range of metal ions with varying oxidation states. However, reports in literature on binding of heparin to metals have investigated only a small sampling of heparin-metal ion interactions. Since interaction of heparin with fluid phase and cell surface macromolecules in vivo is dependent on the heparin structure when bound in a metal ion complex, a survey of the physical parameters for heparin binding to metals is imperative. Atomic absorption and spectrophotometry experiments were performed for metal quantification, and in this study, the relative values for affinity constants and number of binding sites for heparin binding to several alkaline, alkaline earth, main group, and transition metals in their most common oxidation states are reported. We found an overall trend for heparin-metal affinity to be Mn(2+) > Cu(2+) > Ca(2+) > Zn(2+) > Co(2+) > Na(+) > Mg(2+) > Fe(3+) > Ni(2+) > Al(3+)> Sr(2+), with the trend in N (b) being opposite compared with the K (a).