Analysis of Human Blood Plasma Proteome from Ten Healthy Volunteers from Indian Population

Analysis of any mammalian plasma proteome is a challenge, particularly by mass spectrometry, due to the presence of albumin and other abundant proteins which can mask the detection of low abundant proteins. As detection of human plasma proteins is valuable in diagnostics, exploring various workflows with minimal fractionation prior to mass spectral analysis, is required in order to study population diversity involving analysis in a large cohort of samples. Here, we used ‘reference plasma sample’, a pool of plasma from 10 healthy individuals from Indian population in the age group of 25–60 yrs including 5 males and 5 females. The 14 abundant proteins were immunodepleted from plasma and then evaluated by three different workflows for proteome analysis using a nanoflow reverse phase liquid chromatography system coupled to a LTQ Orbitrap Velos mass spectrometer. The analysis of reference plasma sample a) without prefractionation, b) after prefractionation at peptide level by strong cation exchange chromatography and c) after prefractionation at protein level by sodium dodecyl sulfate polyacrylamide gel electrophoresis, led to the identification of 194, 251 and 342 proteins respectively. Together, a comprehensive dataset of 517 unique proteins was achieved from all the three workflows, including 271 proteins with high confidence identified by≥2 unique peptides in any of the workflows or identified by single peptide in any of the two workflows. A total of 70 proteins were common in all the three workflows. Some of the proteins were unique to our study and could be specific to Indian population. The high-confidence dataset obtained from our study may be useful for studying the population diversity, in discovery and validation process for biomarker identification.     

正常人群磁共振全身弥散加权成像特点的初步研究

目的：探讨健康志愿者不同性别各年龄组人体多脏器全身弥散加权成像的图像特征。方法：对50例健康志愿者行全身弥散加权成像扫描,其中,20．30岁、30．40岁、40—50、50．60、60岁以上各二十例,每组内男女各十例。所得图像均经nDeMR后处理工作站（AW4．3）行MIP重建和黑白反转显示技术,并重建ADC图,观察不同性别各年龄组人体多脏器弥散加权图像的特点,并测定不同部位的ADC值,测量的部位主要包括脑灰质、脑白质、尾状核头、丘脑、桥脑、腮腺、肝脏、脾脏、T8椎体、T8—9椎间盘、L2椎体、L2．3椎间盘、前列腺、乳腺、子宫颈等,每个部位重复测量三次取平均值,单位为×10^-3mm^2／S。结果：1、WB-DWI的正常表现。2、不同性别及年龄组间各组织器官ADC值的统计学分析脑白质、T8椎体ADC值在不同性别及年龄组之间存在统计学差异（P〈0．05）;T8．9椎间盘、L2．3椎间盘及女性乳腺ADC值在不同年龄组之间存在统计学差异（P〈0．05）;L2椎体ADC值在男女之间存在统计学差异（P〈0．05）。3、不同部位ADC值平均值。结论：正常人体不同组织器官弥散加权图像特点及ADC值受到性别、年龄的影响,不同性别各年龄组的正常WB-DWI表现存在差异。了解健康成人正常WB-DWI的表现对于准确判断各组织器官的疾病有着重要的意义。     

Alpha-Synuclein Levels in Blood Plasma Decline with Healthy Aging

There is unequivocal evidence that alpha-synuclein plays a pivotal pathophysiological role in neurodegenerative diseases, and in particular in synucleinopathies. These disorders present with a variable extent of cognitive impairment and alpha-synuclein is being explored as a biomarker in CSF, blood serum and plasma. Considering key events of aging that include proteostasis, alpha-synuclein may not only be useful as a marker for differential diagnosis but also for aging per se. To explore this hypothesis, we developed a highly specific ELISA to measure alpha-synuclein. In healthy males plasma alpha-synuclein levels correlated strongly with age, revealing much lower concentrations in older (avg. 58.1 years) compared to younger (avg. 27.6 years) individuals. This difference between the age groups was enhanced after acidification of the plasmas (p<0.0001), possibly reflecting a decrease of alpha-synuclein-antibody complexes or chaperone activity in older individuals. Our results support the concept that alpha-synuclein homeostasis may be impaired early on, possibly due to disturbance of the proteostasis network, a key component of healthy aging. Thus, alpha-synuclein may be a novel biomarker of aging, a factor that should be considered when analyzing its presence in biological specimens.     

Response of Whole Blood,Erythrocyte and Plasma Vitamin E Content to Dietary Vitamin E Intake in the Chick

Whole blood, red blood cells (RBC), and plasma vitamin E (VE) levels in chicks fed dietary VE (dl-α-tocopheryl acetate, dl-αTa) supplementation in steps of 0.0, 5.0, 10.0, 15.0, 20.0 and 30.0 mg/Kg were determined to examine their usefulness as an index of VE status. The increase in VE level was significant and linear in whole blood (r = 0.90), RBC (r = 0.89) and plasma (r = 0.93) in response to dietary VE intake. There was a close correlation between VE in plasma vs whole blood (r = 0.90), plasma vs RBC (r = 0.91) and whole blood vs RBC (r = 0.95). The plasma VE content was 1.2–1.8 times greater than that of whole blood, and 6.6–12.5 times greater than that of RBC. The plasma total lipids content was not affected by the dietary VE intake, whereas the level of VE in the plasma total lipids was significantly increased with increasing supplementation. Alpha tocopherol was the major isomer (ca 92 %) of VE in whole blood, RBC and plasma at hatching. The small proportions of β-tocopherol (ca 2 %), γ-tocopherol (ca 5 %) and α-tocotrienol (ca 1 %) observed at 1 day of age had decreased or totally disappeared by 7 days of age after feeding the VE-free basal diet. The data showed that in the chick, the whole blood and RBC levels of VE were as sensitive and reliable indexes of dietary VE status as was that of the plasma.     

Circadian Variation in the Response to Experimental Endotoxemia and Modulatory Effects of Exogenous Melatonin

Disturbances in circadian rhythms are commonly observed in the development of several medical conditions and may also be involved in the pathophysiology of sepsis. Melatonin, with its antioxidative and anti-inflammatory effects, is known to modulate the response to endotoxemia. In this paper, we investigated the circadian variation with or without melatonin administration in an experimental endotoxemia model based on lipopolysaccharide (LPS). Sixty male Sprague-Dawley rats were assigned to six groups receiving an intraperitoneal injection of either LPS (5?mg/kg), LPS?+?melatonin (1?mg/kg), or LPS?+?melatonin (10?mg/kg) at either daytime or nighttime. Superoxide dismutase (SOD) was analyzed in liver samples collected after decapitation. Furthermore, inflammatory plasma markers (cytokines interleukin [IL]-6, IL-10) and oxidative plasma markers (ascorbic acid [AA], dehydroascorbic acid [DHA], and malondialdehyde [MDA]) were analyzed before and 5?h after the onset of endotoxemia. There were significant higher levels of SOD (p?<?0.05), IL-6 (p?<?0.01), and IL-10 (p?<?0.05) during nighttime endotoxemia compared with daytime. At daytime, melatonin 1 and 10?mg reduced the levels of MDA and increased SOD, IL-6, IL-10, and DHA (p?<?0.05). At nighttime, melatonin reduced the levels of MDA and increased DHA (p?<?0.05). Additionally, 10?mg melatonin resulted in lower levels of AA during daytime (p?<?0.05). No dose relationship of melatonin was observed. The results showed that the response induced by experimental endotoxemia was dependent on time of day. Melatonin administration modulated the inflammatory and oxidative stress responses induced by endotoxemia and also resulted in higher levels of antioxidants during daytime. The effect of circadian time on the endotoxemia response and possible modulatory effects of melatonin need further investigations in a human endotoxemia model.     

Whole Blood Selenium Levels in Healthy Adults from the West of Algeria

The purpose of this study was to assess whole blood selenium levels of 300 healthy adults living in four selected areas of the west of Algeria. Selenium was measured using differential pulse cathodic stripping voltammetry with a detection limit of 29.20 μg/L. The mean of whole blood selenium concentrations was 85.65 ± 21.60 μg/L ranging between 30.90 and 144.04 μg/L. This concentration did not vary significantly (P > 0.05) in relation to the gender of the subject, with concentrations of 87.75 ± 21.30 μg/L in men and 83.95 ± 21.60 μg/L in women group. Individuals older than 60 years had a whole blood selenium concentration significantly lower than the rest of the population. However, the measured selenium concentrations in the residential areas were not statistically different (P > 0.05). A total of 32 (10.70%) individuals exhibited whole blood selenium level below 60 μg/L. These results are similar to those of some European countries but are much lower than data observed in USA or seleniferous regions.     

Circadian and Ultradian Rhythms in Blood Glucose and Plasma Insulin of Healthy Adults

The circadian and ultradian variations of blood glucose and plasma insulin have been characterized individually and as a group phenomenon in five healthy young adults studied while adhering as closely as possible to their usual routine of sleep, activity, meal content and timing. Three complementary methods were used to analyze the data: displaying raw data as a function of time; cosinor method according to Nelson and Halberg; and time series analyses as proposed by De Prins and Malbecq. The subjects were studied in the laboratory and their life routine were controlled, but very close to that of their habitual routine. They had mainly ultradian rhythms of blood glucose (mainly about 6 hr) and circadian rhythms of immunoreactive insulin (I.R.I.). Blood glucose ultradian rhythms seem to be mainly but not exclusively mealtime dependent, while I.R.I, circadian rhythms appear to be primarily endogenous in origin. Therefore, the role played by insulin in the control of blood glucose levels seems to be programmed on a circadian basis rather than by a time independent feedback phenomenon as postulated by the conventional homeostatic hypothesis. The advantage of this chronophysiologic approach is to consider circadian rhythms of both I.R.I. and insulin effectiveness as an adaptive phenomenon able to maintain blood sugar changes in the ultradian domain of rhythms.     

Increased Set Shifting Costs in Fasted Healthy Volunteers

We investigated the impact of temporary food restriction on a set shifting task requiring participants to judge clusters of pictures against a frequently changing rule. 60 healthy female participants underwent two testing sessions: once after fasting for 16 hours and once in a satiated state. Participants also completed a battery of questionnaires (Hospital Anxiety and Depression Scale [HADS]; Persistence, Perseveration and Perfectionism Questionnaire [PPPQ-22]; and Eating Disorders Examination Questionnaire [EDE-Q6]). Set shifting costs were significantly increased after fasting; this effect was independent of self-reported mood and perseveration. Furthermore, higher levels of weight concern predicted a general performance decrement under conditions of fasting. We conclude that relatively short periods of fasting can lead to set shifting impairments. This finding may have relevance to studies of development, individual differences, and the interpretation of psychometric tests. It also could have implications for understanding the etiology and maintenance of eating disorders, in which impaired set shifting has been implicated.     

Changes in Plasma Lipid Levels in Young Healthy Volunteers by Adding an Extruder-cooked Soy Protein to Conventional Meals

The serum cholesterol level in rats was significantly decreased in a group fed on a soyprotein peptic hydrolyzate (SPH) when compared with a group fed on a casein tryptic hydrolyzate (CTH). The fecal excretion of total steroids was significantly greater with rats fed on the SPH diet when compared with the CTH diet. The results of CaCo-2 studies clearly suggest that the suppression of cholesterol absorption in the intestinal epithelia is part of the mechanism for the hypocholesterolemic action induced by SPH.     

Circadian Variation of Fibrinolytic Activity in Blood

Approximately 35 years ago, it was discovered that spontaneous fibrinolytic activity in blood showed a sinusoidal variation with a period of 24 h; it increased severalfold during the day, reaching a peak at 6:OO p.m. and then dropped to trough levels at 3:00–4:00 a.m. The range of the fluctuation and the 24-h mean levels were highly reproducible within an individual; moreover, the timing of the oscillation was remarkably consistent among individuals, with a fixed phase relationship to external clock time. The biorhythm could not be accounted for simply by variations in physical activity, body posture, or sleepfwake schedule. Gender, ethnic origin, meals, or resting levels of blood fibrinolytic activity also did not influence the basic features of the rhythm. Older subjects, compared to younger ones, showed a blunted diurnal increase in fibrinolytic activity in blood. Recent studies have established that, of the known components of the fibrinolytic system, only tissue-type plasminogen activator (tPA) and its fast-acting inhibitor, plasminogen activator inhibitor- 1 (PAL l), show a marked circadian variation in plasma. In contrast, levels of plasminogen, α₂-antiplasmin, urinarytype plasminogen activator, and a reversible tPA inhibitor vary little or none during the 24 h. Quenching antibodies to tPA have shown that the circadian rhythm of fibrinolytic activity in blood is due exclusively to changes in tPA activity. However, the 24-h fluctuation of plasma tPA activity is phase shifted in relation to the rhythm of immunoreactive tPA, but shows a precise phase inversion with respect to the 24-h variation of PAL 1 activity and antigen. Therefore, plasma tPA activity, as currently measured in vitro, is tightly and inversely related to the levels of PAL 1 throughout the 24-h cycle. The factors controlling the rhythmicity of plasma PAI-1 are not fully elucidated but probably involve a humoral mechanism; changes in endothelial function, circulating platelet release. products, corticosteroids, catecholamines, insulin, activated protein C, or hepatic clearance do not appear to be responsible. Shift workers on weekly shift rotations show a disrupted 24-h rhythm of plasma tPA and PAL 1. In acute and chronic diseases, the circadian rhythmicity of fibrinolytic activity may show a variety of alterations, affecting the 24-h mean, the amplitude, or the timing of the fluctuation. It is advisable, therefore, to define the 24-h pattern of plasma tPA and PAI- 1 in patient groups, before levels based on a single blood sampling time are compared to those of a control population. In normal conditions, the 24-h variation of plasma tPA and PAI- 1 is not associated with parallel circadian changes in effective fibrinolysis, assessed as plasma D-dimer concentrations, presumably because fibrin generation in the circulation is low. In diseases in which fibrin formation is increased, however, the physiological drop of fibrinolytic activity in the morning hours may favour thrombus development at this time of day, in agreement with the reported higher morning frequency of acute thrombotic events.     

Erythrocyte Concentrates Recovered from Under-Collected Whole Blood: Experimental and Clinical Results

BackgroundAlthough periodic blood shortages are widespread in major Chinese cities, approximately 1x10⁵ U of whole blood are discarded yearly because of under-collection. To reduce the wastage of acid citrate dextrose solution B (ACD-B) anticoagulated under-collected whole blood (UC-WB), this study was performed to elucidate the effect of extracellular pH and holding time on erythrocyte quality. Mannitol-adenine-phosphate (MAP) erythrocyte concentrates (UC-RBCs) were prepared with UC-WB to assess the safety and efficacy of this component.MethodsThe effect of the different extracellular pH levels and storage times on erythrocytes was assessed by fluorescent probes, SDS-PAGE electrophoresis, electron microscopy and spectroscopy. In vitro properties of 34 UC-RBCs that were prepared with UC-WB at different times after collection were analyzed and compared to normal RBCs during 35 days of storage. The results of transfusion with UC-RBCs and the incidence of adverse reactions in 49 patients were determined.Results1) Low extracellular pH levels and long storage time induced increases in RBC fluorescence polarization and mean microviscosity, changes in membrane fluidity, band 1, 2 and 3 protein expression, and erythrocyte morphology. 2) During storage for 35 days, difference in between-subjects effects of K⁺, hemolysis and supernatant erythrocyte membrane protein (EMP) were statistically significant (P = 0.041, 0.007 and 0.002, respectively), while the differences between these parameters in the 4 h group and comparable controls were less significant. 3) Clinical data from 49 patients confirmed that transfusions with UC-RBCs were satisfactory with no adverse reactions.ConclusionThese results suggest that it is feasible to prepare RBCs with ACD-B anticoagulated UC-WB at a minimum of 66% volume of the labeled collection. It was effective and safe to transfuse the UC-RBCs prepared within 4 h after collection and stored within 7 days. The use of UC-WB would be a welcome addition to limited blood resources in China.

Trial Registration

Chinese Clinical Trial Registry ChiCTR-TRC-13003967     

中国健康老年前期人全血比粘度参考值与地理因素的关系

目的为制定中国健康老年前期人全血比粘度参考值的统一标准提供科学依据。方法收集了全国各地用毛细管法测定的健康老年前期人全血比粘度参考值,并对其与地理因素的关系进行了研究。结果发现海拔高度是影响健康老年前期人全血比粘度参考值最主要的因素,随着海拔高度的逐渐增大,健康老年前期人全血比粘度参考值也在逐渐的增大,相关性很显著。用逐步回归分析的方法推导出了二个回归方程。结论如果知道了中国某地的地理因素,就可以用这二个回归方程估算这个地区的健康老年前期人全血比粘度参考值。依据健康老年前期人全血比粘度参考值与地理因素的依赖关系把中国分为青藏区,西南区,西北区,东南区,华北区,东北区等六个区。     

A Novel Magnetic Stimulator Increases Experimental Pain Tolerance in Healthy Volunteers - A Double-Blind Sham-Controlled Crossover Study

The ‘complex neural pulse’^TM (CNP) is a neuromodulation protocol employing weak pulsed electromagnetic fields (PEMF). A pioneering paper reported an analgesic effect in healthy humans after 30 minutes of CNP-stimulation using three nested whole head coils. We aimed to devise and validate a stimulator with a novel design entailing a multitude of small coils at known anatomical positions on a head cap, to improve applicability. The main hypothesis was that CNP delivery with this novel device would also increase heat pain thresholds. Twenty healthy volunteers were enrolled in this double-blind, sham-controlled, crossover study. Thirty minutes of PEMF (CNP) or sham was applied to the head. After one week the other treatment was given. Before and after each treatment, primary and secondary outcomes were measured. Primary outcome was heat pain threshold (HPT) measured with thermal quantitative sensory testing. Other outcomes were warmth detection threshold, and aspects of cognition, emotion and motor performance. As hypothesized heat pain threshold was significantly increased after the PEMF stimulation. All other outcomes were unaltered by the PEMF but there was a trend level reduction of cognitive performance after PEMF stimulation as measured by the digit-symbol substitution task. Results from this pilot study suggest that our device is able to stimulate the brain and to modulate its function. This is in agreement with previous studies that used similar magnetic field strengths to stimulate the brain. Specifically, pain control may be achieved with PEMF and for this analgesic effect, coil design does not appear to play a dominant role. In addition, the flexible configuration with small coils on a head cap improves clinical applicability.

Trial Registration

Dutch Cochrane Centre NTR1093