Identification of melanoma‐specific serological markers using proteomic analyses

The discovery of novel melanoma markers for not only early detection but also monitoring disease status is promising to improve the clinical outcome of patients. In the present study, we performed proteomic comparative analysis of plasma proteins between healthy volunteers and melanoma patients using NanoLC and MALDI‐TOF‐MS. As a result, we were successful in identifying nine proteins that were specifically expressed in melanoma plasma compared with healthy plasma, most of which had not previously been identified as plasma markers of melanoma. The mRNA expression levels of four proteins [pro‐platelet basic protein precursor (PPBP), serum amyloid A2 (SAA2), complement factor H‐related protein 1 precursor (FHR1), inter‐alpha‐trypsin inhibitor heavy chain H4 precursor (IAIH4)] were prominently up‐regulated in several melanoma cell lines compared with melanocytes. Moreover, two proteins (PPBP, SAA) were shown to be expressed in tumor specimens from melanoma patients. In the survival time analysis regarding melanoma patients, the semi‐quantified plasma PPBP levels were statistically negatively correlated with the survival time. Most interestingly, the significant survival benefit was seen in low PBPP level group (< index 20) versus high level (≥ index 20) group. The results suggest that PPBP might be a novel promising serological marker and a prognostic factor specific to melanomas.     

Analysis of low density lipoprotein apheresis: post-treatment rebound using mathematical models

Double membrane low density lipoprotein (LDL) apheresis (thermofiltration) was applied to treat six hyperlipidaemic patients and five controls 14 and 5 times, respectively, and post-treatment plasma LDL cholesterol (LDLc) concentration rebound was measured. To simulate the rebound, a nonlinear model (NLM) was developed. It consists of two compartments representing plasma and liver. LDLc uptake by the liver is described by receptor-independent and by receptor-mediated transport as controlled by the liver cholesterol concentration. NLM analysis of patient data resulted in pre-treatment fractional catabolic rate (FCR) for LDLc of 0.19 day-1 and the maximum FCR for LDLc of 0.26 day-1 on the 4th day. NLM showed that in patients and controls the post-treatment LDL synthesis rate rose by 6% to 15%. The steady-state (kinetic) version of this model was used to develop a formula describing the relationship between the FCR for LDLc and blood plasma LDLc concentration which agrees well with published data.     

Effectiveness of the Smart Healthcare Glove System for Elderly Persons with Hypertension

With the global population growth of elderly persons, there has been a tendency toward higher quality of life, along with continually increasing expectations and needs for enhanced health care for elderly persons. The purposes of this research were to develop the E‐textile–based Smart Healthcare Glove System (SHGS) with Transcutaneous Electrical Nerve Stimulator (TENS) as a health care device for the elderly hypertensive, and to evaluate the effectiveness of SHGS for hypertensive treatment. The SHGS consists of an inner glove with sensor‐embedded conductive textiles; an outer glove; a TENS for electrical stimulation; and an arm band for a ubiquitous health care wearable device. The SHGS with TENS is used for hypertension treatment and care by electrically stimulating meridian points on the palm. The health care performance test was carried out with a dozen elderly female patients (six Senior‐old‐ageing and six Old‐ageing groups) suffering from hypertension, and was conducted to measure blood pressure and pulse rate before and after wearing the SHGS with TENS for 15 min. Significantly, this study found that the mean blood pressure of all 12 patients after wearing SHGS decreased from 142.58 ± 9.90/82.46 ± 4.45 mmHg to 119.83 ± 9.23/75.79 ± 4.90 mmHg in systolic blood pressure (p < .001) as well as diastolic blood pressure (p < .01) by paired t test. Also, the mean blood pressure of chronic hypertensive patients decreased significantly from 149.25 ± 9.45/82.75 ± 2.70 mmHg to 125.33 ± 9.78/76.67 ± 3.31 mmHg in systolic blood pressure (p < .001). In addition, there was a significant difference in blood pressure before and after wearing the SHGS by Wilcoxon signed rank test (p value: 0.0313, 0.0005). These results suggest that SHGS decreases blood pressure, improves irregular blood circulation, and can be an effective high‐tech health care device for elderly hypertensive patients. Although this study used only elderly participants, the results of the study may be generalized to hypertensive patients at any age. © 2012 Wiley Periodicals, Inc.     

Growth model for cholesterol accumulation in the wall of a simplified 3D geometry of the carotid bifurcation

Atherosclerosis is one of the leading causes of death in the first world countries nowadays. It is a vascular disease that affects medium and large size arteries, involving the formation of plaques within the artery wall. These plaques result from the accumulation of fat, cholesterol, cell debris, smooth muscle cells and other cells and substances, and may cause temporary or definitive lack of blood supply to an organ.This article proposes a model for cholesterol accumulation and plaque growth. The model is basically a mass balance of low density lipoproteins (LDL) in the intima. The inflow, outflow, oxidation, and consumption of LDL is modeled combining partial models and correlations available in the literature.The model was implemented into an open source finite volume code. Assuming steady blood flow, the code was used to predict lesion formation on a three-dimensional model of the carotid artery bifurcation, a location greatly studied for its role in supplying blood to some parts of the brain and for being related to strokes due to formation of atheromas. The simulation was carried out under physiologic conditions for blood pressure and LDL blood concentration.Results for LDL mass accumulation and intimal thickening over time, plaque shape, and location of thicker spots are reported, showing that the proposed model approximates reasonably well the intimal thickening obtained from post-mortem aortic fatty streaks and from B-mode ultrasonography of the carotid artery of healthy subjects reported by other authors.     

Proteins that underlie neoplastic progression of ulcerative colitis

Patients with ulcerative colitis (UC) have an increased risk for developing colorectal cancer. Because UC tumorigenesis is associated with genomic field defects that can extend throughout the entire colon, including the non‐dysplastic mucosa, we hypothesized that the same field defects will include abnormally expressed proteins. Here, we applied proteomics to study the protein expression of UC neoplastic progression. The protein profiles of colonic epithelium were compared with (i) UC patients without dysplasia (non‐progressors), (ii) non‐dysplastic colonic tissue from UC patient with high‐grade dysplasia or cancer (progressors), (iii) high‐grade dysplastic tissue from UC progressors, and (iv) normal colon. We identified differential protein expression associated with UC neoplastic progression. Proteins relating to mitochondria, oxidative activity, and calcium‐binding proteins were some of the interesting classes of these proteins. Network analysis discovered that Sp1 and c‐myc proteins may play roles in UC early and late stages of neoplastic progression, respectively. Two over‐expressed proteins in the non‐dysplastic tissue of UC progressors, carbamoyl‐phosphate synthase 1 and S100P, were further confirmed by immunohistochemistry analysis. Our study provides insight into the molecular events associated with UC neoplastic progression, which could be exploited for the development of protein biomarkers in fields of non‐dysplastic mucosa that identify a patient's risk for UC dysplasia.     

Cell‐Free DNA Blood Collection Tubes Are Appropriate for Clinical Proteomics: A Demonstration in Colorectal Cancer

Background

Optimized blood collection tubes (BCT) have been developed to expand the utility of plasma cell‐free DNA (cfDNA) and are in clinical use. The appropriateness of plasma collected and stored in these tubes for proteomic analysis is unknown.

Methods

Paired blood samples were collected in BCT and traditional K3EDTA (EDTA) tubes from healthy controls and from colorectal cancer (CRC) patients before and after surgery, and stored for between 45 min and 48 h at room temperature. Plasma proteins were analyzed following high‐abundant plasma protein depletion in quantitative discovery and targeted proteomics by liquid chromatography tandem‐mass spectrometry (LC‐MS/MS).

Results

BCT reduced cellular protein contamination in healthy controls over time, and increased the number of high confident low‐abundant protein identifications in CRC blood samples compared to matched samples collected in EDTA tubes. The known CRC plasma protein biomarker, carcinoembryonic antigen (CEA), showed elevated levels across patients pre‐operatively when collected and stored in BCT compared to EDTA tubes. Emerging CRC biomarkers, Dickkopf‐3 (DKK3) and Gelsolin (GSN), showed elevated levels pre‐operatively when collected in BCT.

Conclusions

Optimized BCT are appropriate for low‐abundant plasma protein analysis and can be used with confidence for clinical proteomics.

     

基于BP人工神经网络的胆固醇含量诊断系统

临床研究已证明,血清总胆固醇水平增高是导致冠心病的独立危险因素.血清总胆固醇越高,发生动脉粥样硬化的风险越大,时间也越早.血清总胆固醇每降低1%,发生冠心病的危险性可减少2%.目前一般采用从血样的光谱组成得到血清的胆固醇含量级别.本文使用BP人工神经网络的算法,对几百例血清样本数据进行了训练、确证与测试,结果表明:该方法对胆固醇含量级别HDL、LDL和VLDL的分析取得了较为理想的效果.     

Protein carbonylation in kidney medulla of the spontaneously hypertensive rat

Enhanced generation of ROS has been reported in models of hypertension such as the spontaneously hypertensive rat (SHR). Impairment of kidney function has been implicated in development and progression of hypertension, and the renal medulla appears to play an important role in regulating long‐term blood pressure. A key biomarker of oxidative stress is the formation of protein carbonyls, which we set out to characterize in the SHR medulla. We identified 11 proteins that were differentially carbonylated in SHR medulla in comparison to normotensive wistars including enolase 1, catalase, carbonic anhydrase II, transferrin and members of the aldo–keto‐reductase family. This enhanced protein oxidation was not only accompanied by an increase in intracellular iron deposition, but aldo–keto‐reductase activity was also significantly less in SHR medulla than in normotensive Wistars. Oxidative stress appears selectively to target a subset of proteins in SHR kidney and modification of these proteins may in turn contribute to the renopathy associated with hypertension.     

Proteome profiling of vitreoretinal diseases by cluster analysis

Vitreous samples collected in retinopathic surgeries have diverse properties, making proteomics analysis difficult. We report a cluster analysis to evade this difficulty. Vitreous and subretinal fluid samples were collected from 60 patients during surgical operation of non‐proliferative diabetic retinopathy, proliferative diabetic retinopathy, proliferative vitreoretinopathy, and rhegmatogenous retinal detachment. For controls, we collected vitreous fluid from patients of idiopathic macular hole, epiretinal, and from a healthy postmortem donor. Proteins from these samples were subjected to quantitative proteomics using two‐dimensional gel electrophoresis. We selected 105 proteins robustly expressed among ca. 400 protein spots and subjected them to permutation test. By using permutation test analysis we observed unique variations in the expression of some of these proteins in vitreoretinal diseases when compared to the control and to each other: (i) the levels of inflammation‐associated proteins such as alpha1‐antitrypsin, apolipoprotein A4, albumin, and transferrin were significantly higher in all four types of vitreoretinal diseases, and (ii) each vitreoretinal disease elevated a unique set of proteins, which can be interpreted based on the pathology of retinopathy. Our protocol will be effective for the study of protein expression in other types of clinical samples of diverse properties.     

Computer‐vision determination of 3‐D geometric parameters of LDL particles via cryogenic transmission electron microscopy

Abstract— Previous research has shown that the size of the LDL macromolecules can have an effect on cardiovascular health and that LDL macromolecules may be non‐spherical in shape. Some of these studies, however, used methods that are not conducive to automatic determination of the 3‐D parameters of the particles. In particular, the prior methods used for determination of geometric‐parameter determination were either centrifugal separations or manual determination of parameters from cryogenic transmission electron micrographs. An application of computer‐vision techniques to automatically determine the 3‐D parameters from cryogenic transmission electron microscopy (CTEM) images will be described. Correlation of computer‐generated geometric models to the orthonormal projection CTEM imagery were investigated to determine the applicability of finding the pertinent geometric parameters of the expected discoid shape of the LDL particles. The processing showed that the discoid shape can be verified using small‐angle rotations that are more amenable to the limitations of CTEM imaging.     

Toward the Proteome of the Human Peripheral Blood Eosinophil

Eosinophils (EOSs) are granular leukocytes that have significant roles in many inflammatory and immunoregulatory responses, especially asthma and allergic diseases. We have undertaken a fairly comprehensive proteomic analysis of purified peripheral blood EOSs from normal human donors primarily employing 2‐DE with protein spot identification by MALDI‐MS. Protein subfractionation methods employed included IEF (Zoom^® Fractionator) and subcellular fractionation using differential protein solubilization. We have identified 3141 proteins, which had Mascot expectation scores of 10^?3 or less. Of these 426 were unique and non‐redundant of which 231 were novel proteins not previously reported to occur in EOSs. Ingenuity Pathway Analysis showed that some 70% of the non‐redundant proteins could be subdivided into categories that are clearly related to currently known EOS biological activities. Cytoskeletal and associated proteins predominated among the proteins identified. Extensive protein posttranslational modifications were evident, many of which have not been previously reported that reflected the dynamic character of the EOS. This data set of eosinophilic proteins will prove valuable in comparative studies of disease versus normal states and for studies of gender differences and polymorphic variation among individuals.     

Proteome analysis of a plasma membrane-enriched fraction at the placental feto-maternal barrier

Purpose : We want to identify proteins that are part of or associated with the plasma membrane of the human feto‐maternal barrier, which is crucially important for nutrient, gas, and waste exchange between the mother and the fetus. All transfer processes occur through one specialized endothelial cell layer, the multinuclear syncytiotrophoblast (STB). Specifically, the apical plasma membrane of the STB interacts with the maternal blood and is the site of initial transport processes across the placenta. Experimental design : We used a proteomic approach that employed the enrichment of apical STB membranes isolated from healthy placentae by ultracentrifugation and saccharose gradient centrifugation steps in combination with 1‐D SDS‐PAGE and ESI‐MS analysis. Results : We identified 296 different proteins, 175 of which were integral and peripheral membrane proteins, partially containing 1–12 transmembrane domains or lipid anchors. One hundred and sixty‐one proteins (54%) were allocated to the plasma membrane. Conclusions and clinical relevance : A high number of transporters, receptors, and proteins involved in signal transduction processes and vesicular trafficking were identified for the first time at the feto‐maternal barrier. Our results are valuable sources for further studies of the cell physiology of the healthy placenta at the time of birth or the pathophysiology of several pregnancy disorders.     

脉搏传播时间与血压关系的长时记忆性分析

相对于柯氏音法,通过脉搏传播时间估算血压不仅更为便携,还可以实现血压的连续测量。但是因为现有研究建立的线性方程的有效时间较短,所以脉搏传播时间随血压变化的机制有待进一步的分析。文中以MIMIC数据库中的10例数据为研究对象,从长时记忆的角度,以符号化和复杂网络为主要研究手段分析了血压与脉搏传播时间的关系。对网络的度分布进行了分析,结果显示收缩压网络度分布具有幂率性,验证了收缩压脉搏波传播时间关系序列的长时记忆。对血压网络节点变化的分析显示,相对于舒张压,收缩压网络的节点数能较快达到饱和,反映了某种核心状态对血压脉搏传播时间关系的持续影响。研究结果可以为通过脉搏波传播时间更精确地无创连续测量血压提供支持。     

Thymosin β4 is differentially expressed in the cerebrospinal fluid of Creutzfeldt‐Jakob disease patients: a MALDI‐TOF MS protein profiling study

MALDI‐TOF protein profiling analysis permits the detection of peptides and small proteins in complex protein mixtures with great accuracy. We applied this analysis to cerebrospinal fluid (CSF) from 15 patients affected by Creutzfeldt‐Jakob disease (CJD). We compared the levels of the normalized ion signals of 11 sporadic and 4 genetic CJD forms with those from ten healthy control subjects and eight non‐CJD relapsing‐remitting multiple sclerosis patients. In so doing, we detected 61 differentially expressed ion signals in CJD samples compared to controls. Among the 61 signals, 3 signals had significantly increased levels with high statistical significance (p <0.0001) and were located at 3238.3 m/z, 4963.7 m/z, and 8565.3 m/z. We characterized the 5.0 and 8.6 kDa proteins as thymosin β4 N‐acetylated and free ubiquitin, respectively, while the 3.2‐kDa peptide remained uncharacterized. Although elevated ubiquitin levels have previously been described in CJD, we have demonstrated for the first time the involvement of thymosin β4 in a neurodegenerative disease. To support the validity of thymosin β4 levels obtained by MALDI‐TOF analysis, an independent enzyme immunoassay analysis was performed. Moreover, a validation cohort consisting of CSF from three CJD patients, five healthy subjects, and six non‐CJD relapsing‐remitting multiple sclerosis patients was analyzed in a similar way, yielding superimposable results. We propose that thymosin β4 is a potential new candidate marker for the ante mortem diagnosis of CJD disease.     

Plasma proteome of buffaloes

The proteomic approach has aroused the interest of veterinary medicine researchers, especially regarding the production of biopharmaceuticals and diagnosis of diseases in farm animals. Water buffaloes have gained prominence in the world economy due to the quality of their milk, meat, and leather, in addition to being an important donor of blood components. This work aimed to identify and characterize the proteins present in the blood plasma of Murrah buffaloes (Bubalus bubalis) through 2D electrophoresis, in gel protein digestion followed by mass spectrometry technique and for albumin depletion, in solution protein digestion followed by shotgun analysis. Our results showed the identification of 112 protein spots and 35 individual proteins, respectively. The abundant proteins were represented by albumin, fibrinogen‐α, fibrinogen‐β, fibrinogen‐γ, immunoglobulins in general, α‐1‐antiproteinase, α‐1B‐glycoprotein, α‐2‐HS‐glycoprotein, α‐macroglobulin, apolipoprotein A1, antithrombin‐III, endopin 2B, fetuin‐B, retinol‐binding protein, serotransferrin, transthyretin and vitamin D‐binding protein. Most of these proteins are related to the signaling pathways of the complement system and coagulation cascade. The results allowed a better understanding of the protein composition of these blood components, thus promoting studies on animal health in the search for molecular markers of zoonotic diseases in buffaloes.     

Differential valine metabolism in adipose tissue of low and high fat-oxidizing obese subjects

Differences in fat metabolism are of importance in relation to energy balance. Low fat‐oxidizers (LFO) are thought to be more prone for developing obesity. We studied whether LFO have different fasting adipose tissue (AT) protein profiles than high fat‐oxidizers (HFO). Six LFO and six HFO subjects were selected from an obese group (n = 99, body mass index>30 kg/m²) taking part in a multi‐center study (Nutrient‐Gene interaction in human obesity) based on the postprandial fat oxidation capacity after a high fat load. AT protein profiles were studied by 2‐DE. Differential proteins were clustered with MAPPfinder according to their function. Protein profiles of purified blood cells and adipocytes served to confine the comparison to adipocyte‐specific proteins in AT profiles of LFO and HFO subjects. LFO had increased mitochondrial ROS scavengers possibly related to long‐chain unsaturated fatty acid‐induced increases in mitochondrial ROS‐production. Carbohydrate oxidation seemed to be reduced since expression of several proteins from the glycolysis pathway was lower in LFO. Up‐regulation of the valine catabolism at the level of methylmalonate‐semialdehyde dehydrogenase appeared to be (part of) the compensatory mechanism. In conclusion, the fasting AT protein profile of LFO and HFO differ at the level of ROS scavenging, the glycolysis pathway and valine metabolism.     

Carbon‐nanotube electron emitters for display applications

Abstract— The optoelectronic, mechanical, and thermal properties of carbon nanotubes have made them very attractive for a wide range of potential applications. However, many applications require the growth of aligned/micropatterned carbon nanotubes with or without a modified nanotube surface. We have developed several simple pyrolytic methods for large‐scale production of aligned carbon‐nanotube arrays perpendicular to the substrate surface. We have also used photolithographic and soft‐lithographic techniques to pattern our aligned carbon nanotubes with submicron resolution. These aligned carbon‐nanotube arrays can be transferred onto various substrates of particular interest (e.g., on conducting substrates as electron emitters for flat‐panel displays) in either a patterned or non‐patterned fashion. The well‐aligned structure further allows us to prepare aligned coaxial nanowires of carbon nanotubes sheathed with polymers and to modify the surface of individual carbon nanotubes by plasma treatment. These aligned/micropatterned carbon nanotubes with and without surface modification possess desirable properties for electron emission applications.     

STM32的无创连续血压测量系统设计

针对目前临床对血压的连续测量主要采用有创的方法,本文设计通过脉搏波传导时间(PWTT,Pulse Wave Transmit Time)实现无创连续测量血压的系统.该系统通过采集光电容积脉搏波(PPG)信号和心电(ECG)信号,利用STM32控制芯片将两者信号融合,再通过算法求出特征点,拟合方程求出个体的血压.通过此方法有效实现了无创连续的血压测量.     

The murine plasma protein response to polymicrobial intra-abdominal sepsis

Protein biomarkers in the peripheral blood could potentially be used as early indicators of sepsis and a means to stratify patients for clinical trials. Although individual molecular markers have been proposed for sepsis, none has clinical utility. The global changes in plasma proteins over the clinical course of sepsis have not been characterized using proteomic methods. We used cecal ligation and puncture to induce polymicrobial sepsis in mice and generated plasma protein profiles using 2‐D DIGE of plasma from septic mice and surgical controls. Replicate cohorts (n = 3) of 4–7 animals each were used to identify 62 gel features that changed significantly (Student's t‐test, p<0.05). We identified a suite of plasma proteins that describe uniquely the host plasma response to polymicrobial septic insult. Principal components analysis of protein abundance showed that ～90% of the variability between samples was due to sepsis. In addition to canonical acute phase proteins, we identified proteins that are associated with metabolic changes (e.g. α‐2 HS glycoprotein and zinc α‐2 glycoprotein) consistent with the pathophysiology of sepsis. The panel of sepsis‐associated molecular markers identified herein may prove useful in the diagnosis and categorization of sepsis.