Evaluation of different methods used to measure malonyldialdehyde in human erythrocytes

Malonyldialdehyde (MDA), a secondary product of lipid peroxidation is widely used as an indicator of lipid peroxidation. Eight of the most frequently used methods for measuring MDA have been evaluated with regard to their sensitivity and reproducibility. The sensitivity of these methods for pure MDA solution was in the order: Satoh's > Stocks and Dormandy's > Buege and Aust's > Dresel's > Slater's > Yoshioka's et al. > Yagi's > Jain's method. Whereas the sensitivity of the first four methods for erythrocyte MDA was in the order: Stocks and Dormandy's > Buege and Aust's > Satoh's > Dresel's method. The reproducibility (expressed as coefficient of variation) of these four methods for erythrocyte MDA were: 3.5%, 17.9%, 31.5% and 16.1%, respectively. These results indicate that Stocks and Dormandy's method has the highest sensitivity and an excellent reproducibility for erythrocyte MDA. Also, it was found to be simple and many samples can be treated in a relatively short time. When standard MDA (0.1-15 nmol/ml) was incubated with erythrocytes, the percentage recovery of MDA (using Stocks and Dormandy's method) has ranged from 50-85%. This result indicates that a considerable amount of MDA formed in erythrocytes probably reacts with other cell components and becomes undetectable. Despite this, the determination of MDA level remains a useful indicator of lipid peroxidation and correlates well with the degree of oxidative stress.     

Reproducibility of brachial ultrasonography and flow-mediated dilatation (FMD) for assessing endothelial function

Abstract Background : High-resolution brachial artery ultrasonography is used to study vasodilator response induced by physiologic reactive hyperaemia. We examined the reproducibility of measuring flow-mediated dilatation (FMD) on two occasions.
Aims : To determine the degree of variability of this technique in our vascular laboratory for the design of clinical research studies.
Methods : Nineteen subjects were studied on two separate occasions using an Acuson 128 ultrasound device and a 7.0 MHz linear array transducer. Reactive hyperaemia was induced in the brachial artery by inflation and release of a blood pressure cuff. Nitrate-induced dilatation was assessed in 11 of the 19 subjects. Measurements were made by two observers blinded to subject details.
Results : The 11 subjects given sublingual GTN during the first ultrasound study had a mean nitrate-induced dilatation of 20.7% (sd 9.6). The mean vessel diameter of 3.78 mm (sd 0.7) at rest and 3.89 mm (sd 0.7) during reactive hyperaemia yielded a mean FMD of only 3.0% (sd 2.7). The mean difference in FMD within-observers was 0.13% (sd 2.07), between-observers 0.06% (sd 2.17) and between-studies was 0.57% (sd 6.83).
Conclusions : The reproducibility of FMD measured by brachial artery ultrasound was poor and likely to render the measurements inaccurate for clinical research in our hands. Between-study variation contributed the largest proportion of total study variability. We suggest that investigators using this technique conduct their own careful reproducibility studies in order to avoid the misinterpretation of 'negative' studies.     

Response to aspirin and clopidogrel monitored with different platelet function methods

Laboratory non-response to aspirin or clopidogrel is defined as an inability to cause in vitro detectable platelet function inhibition. It would be beneficial to monitor response to aspirin or clopidogrel with widely available and routinely used platelet function methods, like the platelet function analyzer (PFA-100) or the fully automated coagulation analyzer BCT. The aim of this study was to assess the potential of the coagulation analyzer BCT and the platelet function analyzer PFA-100 in monitoring the response of aspirin and clopidogrel. A group of 125 consecutive patients with arterial occlusive disease treated either with aspirin 100 mg/day (82 patients) or clopidogrel 75 mg/day (43 patients) as only antiplatelet drug were investigated. For the first time platelet-enriched plasma (PRP), not adjusted to a fixed predetermined concentration of platelets, was used for aggregation studies and the effect of clopidogrel alone without combination of aspirin treatment on platelet function was investigated. Response to aspirin was observed in 85% (70/82) of patients using PFA-100, while performing the arachidonic acid-induced aggregation on the BCT showed an inhibitory effect to aspirin in 91% (75/82) of patients. Non-response to aspirin was assessed with both platelet function methods in 7% (6/82) of patients. Clopidogrel response was observed in 58% (25/43) of patients when performing ADP-induced aggregation on the BCT. On the PFA-100 the antiplatelet effect of clopidogrel could not be detected. In conclusion, measurement of platelet aggregation on the BCT using native platelet-enriched plasma allows the quantification of individual inhibitory effects to aspirin as well as to clopidogrel, while the PFA-100 seems only suitable to investigate the degree of platelet inhibition induced by aspirin but not by clopidogrel.     

Flow-mediated changes in pulse wave velocity: a new clinical measure of endothelial function.

AIMS: To test whether measuring hyperaemic changes in pulse wave velocity (PWV) could be used as a new method of assessing endothelial function for use in clinical practice. METHODS AND RESULTS: Flow-mediated changes in vascular tone may be used to assess endothelial function and may be induced by distal hyperaemia, while endothelium-mediated changes in vascular tone can influence PWV. These three known principles were combined to provide and test a novel method of measuring endothelial function by the acute effects of distal hyperaemia on upper and lower limb PWV (measured by a recently developed method). Flow-mediated changes in upper and lower limb PWV were compared in 17 healthy subjects and seven patients with stable chronic heart failure (CHF), as a condition where endothelial function is impaired but endothelium-independent dilator responses are retained. Corroborative measurements of PWV and brachial artery diameter responses to endothelium-dependent and -independent pharmacological stimuli were performed in a further eight healthy subjects. Flow-mediated reduction of PWV (by 14% with no change in blood pressure) was found in normal subjects but was almost abolished in patients with CHF. PWV responses appear to be inversely related to and relatively greater than brachial artery diameter responses. CONCLUSION: The method may offer potential advantages of practical use and sensitivity over conduit artery diameter responses to measure endothelial dysfunction.     

Relation of depression to coronary endothelial function

The purpose of this study was to determine the association between depression and coronary endothelial function and cardiac risk factors in men and women without obstructive coronary artery disease. Patients with no significant coronary artery disease who underwent invasive coronary endothelial function assessment with acetylcholine were studied. Men and women were divided into 2 groups: those with depression and those without. Endothelial function and risk factor profiles were compared between the 2 groups. Seven hundred fifty-nine patients were studied, 603 (79%) without depression and 156 (21%) with depression. Patients with depression were more likely to be women (71% vs 60%, p = 0.02), have greater body mass indexes (29.9 +/- 6.7 vs 28.2 +/- 5.9 kg/m(2), p = 0.002), and be diabetic (12% vs 6%, p = 0.02). Depressed patients also had higher levels of C-reactive protein (0.35 vs 0.30 mg/dl, p = 0.02). There was no difference in the change in coronary blood flow or diameter in response to acetylcholine between the 2 groups in men and women. In conclusion, the results of this study demonstrate that depression is not associated with coronary endothelial dysfunction in men and women without significant coronary artery disease. It is, however, associated with a cluster of cardiac risk factors that are linked to the progression of atherosclerotic disease.     

Hyposplenism: comparison of different methods for determining splenic function

Asplenic patients are at risk for pneumococcal sepsis. Patients with hyposplenic function, such as associated with sickle cell disease (SCD), are also at risk. However, tests to assess splenic function are either unavailable or lacking standardization. The aim of this study was to compare different methods for determining splenic function. Eighteen patients with SCD (i.e., 10 heterozygous (SC) and 8 homozygous (SS) SCD patients), and eight splenectomized patients were compared to 10 controls. All subjects underwent spleen scintigraphy, after which functional splenic volumes (FSV) were calculated. FSV was compared to immunological function and B cell-subsets, as well as phagocytic function represented by the presence of Howell Jolly bodies (HJB) and percentages of pitted red cells (PIT). Heterozygous SCD (SC) patients had increased splenic volumes, but diminished FSV, homozygous SCD (SS) patients were asplenic. Splenectomized and SS patients had a strongly reduced phagocytic and immunological function. SC patients had reduced anti-polysaccharide responses without an increase in PIT. FSV correlated significantly with phagocytic and immunological function. HJB were indicative of splenic dysfunction, HJB absence was not indicative of normal functioning splenic tissue. Although visualizing HJB is methodologically advantageous to PIT, both are valid biomarkers of splenic dysfunction. The amount of non-switched memory B cells is strongly correlated to FSV.     

Reproducibility of ventricular function measurements by contrast angiography

Interobserver reproducibility of single plane contrast angiographic measurements of end-diastolic volume (EDV/m2), end-systolic volume (ESV/m2), stroke volume (SV/m2), and ejection fraction (EF) was studied in two groups of patients. The first group (n = 42) was an unselected, consecutive series of patients in which the calculations of volume were performed manually, representing day-to-day results from a clinical quantitative angiographic laboratory. The second group (n = 31) was selected on the basis of optimal filming technique and volume calculations were performed digitally with the aid of a computer. This group characterizes the reproducibility that could be attained for investigational purposes. In the selected group, interobserver reproducibility of four regional ejection fractions was determined. The correlation coefficient (r), standard error of the estimate (Sy.x), average difference, and 95% confidence limits are given. Measurement variation accounted for 9-15% of the variance in this study for EDV/m2 (consecutive series), indicating that caution is needed when relating contrast angiographic measurements to other (e.g., noninvasive) measures of volume by simple linear regression.     

A cross-over evaluation of different methods and devices to measure blood pressure in type 1 diabetic patients with nephropathy

BACKGROUND: In type 1 diabetic patients with nephropathy, tight blood pressure control has been shown to prevent the progression of the disease. Up until now, self-monitoring, ambulatory and office blood pressure values have not been compared in these patients. Thus, we have evaluated blood pressure values obtained in the office by a physician and at home by self-monitoring with those measured under ambulatory conditions in these patients. Additionally, for blood pressure self-monitoring, three different devices (the sphygmomanometer, upper-arm oscillometer and wrist oscillometer) were compared. METHODS: Twenty-one treated hypertensive type 1 diabetic patients [age 45+/-9 years, duration of diabetes 33+/-12 years (mean+/-SD)] with overt diabetic nephropathy participated in this study. At both baseline and the end of the study, daytime ambulatory blood pressure measurement was performed. Office blood pressure was measured at baseline. Additionally, all the patients measured their blood pressure over a 3-week period using each of the three different devices, in random order, for 1 week. RESULTS: The mean office blood pressure values (135+/-21/85+/-12mmHg) were higher than both the ambulatory (131+/-23/80+/-12, P<0.05) and self-monitoring values (130+/-14/78+/-10; P<0.05 for systolic and P<0.02 for diastolic values). The difference between the ambulatory and self-monitoring values were not statistically significant. Diastolic blood pressure values measured with the oscillometric wrist device showed a trend towards higher values when compared to those measured with the sphygmomanometer and with the oscillometric upper-arm device (P=0.065 for diastolic values). CONCLUSION: Office blood pressure measurements may over-estimate blood pressure in patients with type 1 diabetes and diabetic nephropathy. Because the oscillometric wrist device tends to over-estimate diastolic values, upper-arm devices should be preferred for blood pressure self-monitoring in these patients.     

Reproducibility and sensitivity to change of four scoring methods for the radiological assessment of osteoarthritis of the hand

BACKGROUND: Osteoarthritis (OA) of the hand could be a relevant model to study the progression of OA in structure-modification trials. Various methods are proposed to assess hand OA and its progression radiologically. OBJECTIVE: To compare intra-reader and inter-reader precision and sensitivity to change of four radiological scoring methods proposed in hand OA. METHODS: 2 trained readers scored separately 105 pairs of radiographs (baseline; year 1), selected from patients enrolled in a randomised controlled trial, for inter-reader reliability and sensitivity to change. They scored twice 60 pairs among the 105 for cross-sectional and longitudinal intra-reader reliability. Radiological hand OA assessment used: global, Kellgren-Lawrence (KL), Kallman and Verbruggen scoring methods. Inter- and intra-reader reliabilities were studied using intraclass coefficient (ICC) and the Bland-Altman method. Sensitivity to change was compared by calculating the standardised response means. RESULTS: Transversal intra-reader reproducibility ICCs ranged from 0.922 to 0.999. Verbruggen ranked the highest, followed by the KL and Kallman methods. Inter-rater reliability was higher for the Verbruggen scores, followed by the KL, global and Kallman scores (ICC 0.706-0.999). Longitudinal intra-reader reliability (baseline; year 1) was better using the Kallman and KL (ICC 0.986 and 0.990), followed by the Verbruggen (0.941) or global methods (0.939). Standardised response means ranged from 0.24 (KL) to 0.29 (Kallman). CONCLUSION: All four methods compared well with respect to reliabilities. However, the Verbruggen and Kallman methods performed better. The method most sensitive to change was the Kallman method, followed by Verbruggen and global scores. This study also suggests that structural changes could be detected in hand OA over a 1-year period.     

Effect of different anesthesia methods on erythrocyte immune function in mice

ObjectiveTo explore effect of different anesthesia methods and different anesthetics on erythrocyte immune function in mice.MethodsThe mice were anesthetized by isoflurane and ether inhalation, and also under intraperitoneal anesthesia with sodium pentobarbital and chloral hydrate. Blood was collected from the ventro-cardinal vein. Automatic blood cell analyzer was used for routine blood examination, and the canthine oxidase method was used to measure the superoxide dismutase (SOD) activity. Lipid peroxidation product malondialdehyde (MDA) was measured with TBA, and glutathione peroxidase (GSH-Px) was measured with DTNB, and then the effect of different anesthesia methods and different anesthetics on erythrocyte immune function in mice was observed.ResultsHct level of chloral hydrate intraperitoneal injection group was significantly higher than the other three groups (P<0.05). And the MDA levels in the pentobarbital sodium group were significantly higher than the other three groups (P<0.05). SOD and GSH-Px of the chloral hydrate and sodium pentobarbital intraperitoneal injection group were significantly lower than the other two groups; RBC-C 3bRR and RBC-ICR of the chloral hydrate and sodium pentobarbital intraperitoneal injection group were significantly lower than the other two groups.ConclusionsDifferent drugs can induce changes in immune function of mice at different levels. Isoflurane and ether have less damage to animal body, while chloral hydrate and sodium pentobarbital intraperitoneal injection have a certain inhibitory effect on the animal body respiratory system and can cause greater damage to the body. Therefore, the reasonable selection and control of anesthetics are very important in order to avoid the experimental errors caused by anesthesia.     

Noninvasive methods to measure airway inflammation: future considerations.

This last contribution to the series focuses on open questions regarding: 1) methodological issues; and 2) the potential clinical application of the noninvasive methods such as induced sputum and the analysis of exhaled air for the assessment of airway inflammation. In addition their potential future role in occupational health and the early diagnosis of neoplastic lesions are briefly discussed. The future clinical application of noninvasive methods will depend on the progress made to improve their practicability, particularly in rendering them less time consuming and cheaper. To assess their clinical value, prospective studies are needed to establish whether patients actually benefit from the results obtained. This is also important to implement the methods into the healthcare system and to obtain adequate financial compensation. Therefore, it is necessary to know: 1) whether the assessment of airwav inflammation can aid in coming to an earlier and better defined diagnosis; 2) whether by repeated monitoring it is possible to avoid exacerbations through earlier interventions; and 3) whether the long-term outcome of patients is improved through knowledge of the type and degree of airway inflammation that is taken into account in selecting the appropriate treatment. In the meantime a wealth of data has become available, both for induced sputum and the analysis of exhaled air, which give these methods the potential to be incorporated into future clinical practice. This, however, will, amongst the other issues, depend on favourable cost-benefit ratios which should also be the subject of future prospective studies.     

Measuring endothelial function

Dysfunction of the endothelium and of the arterial wall is well described in patients with atherosclerosis, diabetes, and other risk factors for vascular disease. In recent years, clinical research has focused on elucidating the role of this dysfunction in influencing vascular disease progression. Alteration in the structure of arteries and disruption of the homeostatic functions of the endothelium act as a substrate for end-organ damage and the occurrence of vascular events. Dysfunction of the vascular endothelial cells is probably the earliest event promoting atherosclerotic lesion formation. Therefore, methods capable of assessing endothelial function at a preclinical stage hold potential to refine cardiovascular risk stratification and serve as a guide to monitor the effects of therapeutic interventions. A number of methodologies are currently employed to assess endothelial function, but the optimal approach is not firmly established. In this article, we critically appraise the use of different methodologies employed to study endothelial function as a surrogate marker of future cardiovascular risk.     

不同糖耐量人群血管内皮功能及可溶性选择素P的变化

对33例NGT、30例IGT和30例T2DM应用超声测量其肱动脉内径（DBA），计算收缩期血压由200mmHg降为0时DBA增大所代表的内皮依赖性血管舒张功能（EDF）；测定可溶性选择素P和其他指标。结果显示：（1）EDF值，T2DM组低于IGT和NGT组，IGT组低于NGT组，差异有统计学意义。（2）IGT与T2DM组均显示：EDF值与HOMA-IR及可溶性选择素P呈显著负相关。     

Reproducibility of arterial stiffness indices in different vascular territories and between different observers

Background: Increased arterial stiffness (AS) corresponds to an increase in cardiovascular risk. According to recent guidelines AS parameters can be measured on all superficial arteries. Objective: Proceeding from the assumption that viscoelastic properties differ along the arterial tree we set ourselves the task to study the reproducibility of AS indices measured at the common carotid, brachial and femoral arteries. Methods: The initial study population included 75 patients (40 ± 14.5 years, 45% males) with a variable distribution of cardiovascular risk factors and without clinical evidence of coronary artery disease. AS parameters were measured at the common carotid, brachial and femoral arteries in all patients using echo‐tracking (ET) technique. In a subgroup of 36 patients we tested the interobserver variability in the three vascular territories. Results: We found that there was a significant correlation between AS indices measured at the common carotid and femoral artery (with the only exception for augmentation index) and that AS parameters measured at the brachial artery did correlate neither with common carotid nor with femoral artery indices. The interobserver variability of ET derived AS parameters was good when they were measured at the carotid or femoral artery. The values of AS indices at the brachial artery however showed considerably lower interobserver agreement. Conclusion: The reproducibility of ET derived AS parameters was good when AS was measured at the common carotid or femoral arteries. On the basis of our results brachial artery is probably not a reliable site for AS measurement. (Echocardiography 2011;28:448‐456)     

改善高血压患者血管内皮功能的药物治疗进展

高血压病是严重威胁人类健康和生命的常见疾病。有研究证实人类的高血压患者存在血管内皮功能减退,后者又加速了高血压靶器官的损害。目前认为血管内皮细胞所分泌的一氧化氮(NO)/内皮素(ET-1)的失衡在高血压的发生、发展中起重要作用。内皮细胞是"内皮-高血压-心血管事件"链的始动因子和载体[1]。