共查询到20条相似文献,搜索用时 46 毫秒
1.
Pakharukova NA Pastushkova LKh Trifonova OP Moshkovskiĭ SA Larina IM 《Fiziologiia cheloveka》2011,37(2):77-85
For analysis of inter-individual variability in low-molecular serum subproteome proteome profiles of healthy men at the age of 20-30 years (36 subjects), 30-40 years (11 subjects) and 40-50 years (11 subjects) were obtained. Serum samples were fractionated on magnetic beads MB WCX using ClinProt robot prior to mass-spectrometry based profiling. Mass-spectra were obtained with time-of-flight mass-spectrometer Autoflex III ("Bruker Daltonics") in automatic mode. It was shown that low-molecular serum subproteome of healthy humans was characterized by significant inter-individual variability. 21% of all peaks in proteome profiles had coefficient of variation more than 50% and 29% of all peaks had low dispersion (CV < 30%).Therefore majority of peaks in proteome profile were peaks with moderate inter-individual variability (CV from 30% to 50%). Fragments of high-molecular kininogen, inter-alpha-trypsin inhibitor, complement components C3 and C4a, apolipoprotein CI, platelet factor IV, beta2-microglobulin and cystatin C showed wide variation among examined groups of healthy men. Dispersion of high-molecular kininogen, inter-alpha-trypsin inhibitor, apolipoproteins AII and CIII peaks increased with age. 相似文献
2.
N. A. Pakharukova L. Kh. Pastushkova S. A. Moshkovskii I. M. Larina 《Biochemistry (Moscow) Supplemental Series B: Biomedical Chemistry》2011,5(3):203-212
The aim of this review is to analyze results of studies on characteristics of protein variability and diversity of posttranslational
modifications of proteins in healthy humans. Numerous studies have demonstrated that a proteomic profile is characterized
by significant intra- and inter-individual variability, and quite often natural (“normal”) variability of some proteins can
be comparable to changes observed in pathological processes. Results obtained by our research group have demonstrated high
intra-individual variability of serum low-molecular subproteome of healthy volunteers, certified by a special medial committee
(the coefficient of variation (CV) of 42.6%). The proteins characterized by high variability under normal conditions (e.g.
haptoglobin — 0–40 mg/ml; lysozyme — 0.01–0.1 mg/ml; C-reactive protein — 0.01–0.3 mg/ml) cannot be considered as potential
biomarkers of diseases. On the contrary, proteins and peptides characterized by insignificant dispersion in healthy population
(such as albumin ( CV = 9%); transferrin-(CV = 14%); C3c complement (CV = 17%), α-1 acid glycoprotein (CV = 21%), α-2-macroglobulin
(CV = 20%); transthyretin fragment (CV = 28.3%) and α2-HS-glycoprotein βchain (CV = 29.7%)) can provide valuable information
about the state of health. Thus, studies of plasticity in the proteomic profiles of healthy humans will help to correct reference
intervals used in clinical proteomics. 相似文献
3.
Ermakova II Sakuta GA Potekhina MA Fedorova MA Hoffmann R Morozov VI 《Biochemistry. Biokhimii?a》2011,76(3):359-365
The major proteoglycans from L6J1 rat myoblast culture were identified. The proteoglycans were isolated from different constituents
of cell culture: culture medium, extracellular matrix (ECM), and myoblasts. To identify their core proteins, the proteoglycans
were treated with enzymes specifically digesting chondroitin/dermatan sulfates or chondroitin sulfates. Subsequent electrophoresis
and mass spectrometry revealed versican, collagen XII, and inter-α-trypsin inhibitor classified as chondroitin sulfate proteoglycans
and biglycan known to be chondroitin/dermatan sulfate proteoglycan. Versican was identified in ECM and the other proteoglycans
in the culture medium. Such difference in localization is likely to be a consequence of different biological functions. Versican,
collagen XII, and biglycan are synthesized by myoblasts and inter-α-trypsin inhibitor originates from fetal bovine serum (a
culture medium component). 相似文献
4.
Summary The presence of protein HC-(α1-microglobulin) and inter-α-trypsin inhibitor was investigated in different human tissues. Inter-α-trypsin inhibitor is a
complex protein composed of bikunin and two heavy polypeptide chains. Protein HC and bikunin are transcribed from a common
gene. Inter-α-trypsin inhibitor immunoreactivity was detected in mast cells. The positive reaction could be blocked by antisera
absorption with bikunin, indicating that mast cells contain only bikunin. Protein HC immunoreactivity was revealed on elastic
fibres in connective tissue of skin, colon and lung, and on the internal elastic lamina of blood vessels. In the testis, the
basement membrane of the seminiferous tubules reacted positively with protein HC antibodies. 相似文献
5.
Serum samples from four European populations — Germans from Jena, Germans from Hannover, Italians from Udine, and Slovakians
from Bratislava — were tested for the inter-α-trypsin inhibitor (ITI) polymorphism. The results are compared with those reported
for other European, Asiatic and African populations. Reviewing all the hitherto published ITI allele frequencies one can see
that the variability among European Populations seems to be rather small, whereas Asiatic and African populations are striking
by quite different distribution patterns. 相似文献
6.
O. A. Valeeva L. Kh. Pastushkova N. A. Pakharukova I. V. Dobrokhotov I. M. Larina 《Human physiology》2011,37(3):351-354
The protein composition (proteome) of the body fluids is rather flexible; it can change, responding to various factors of
the external environment and changes in the internal environment. In order to study the variability of the proteome profile
in healthy humans under the conditions of total control of vital rhythm, physical activity, and diet, urine samples were collected
from subjects who had been selected according to special criteria and qualified as healthy by a special physical evaluation
board. The subjects took part in an experiment with a 105-day-long isolation in a pressurized compartment, carried out by
using an autonomous life-support system at the Institute of Biomedical Problems, Russian Academy of Sciences. The purification
and concentration of proteins from the urine samples were carried out using a MB-HIC C8 magnetic bead set (Bruker Daltonics).
The mass spectra have been obtained using an Autoflex III time-of-flight mass spectrometer (Bruker Daltonics) in the positive
lineal mode. One hundred and seventeen peaks were obtained for each urine sample; technological errors of the method have
been studied. The high variability of the urine proteome profile (36 protein MC peaks on average) was shown in healthy humans
under the conditions of isolation and controlled vital activity. 相似文献
7.
A. T. Kopylov E. V. Ilgisonis O. V. Tikhonova T. E. Farafonova S. E. Novikova V. G. Zgoda E. A. Ponomarenko A. V. Lisitsa A. A. Markin B. V. Morukov A. I. Grigoriev A. I. Archakov 《Doklady. Biochemistry and biophysics》2017,476(1):326-328
Quantitative proteomic analysis of 50 blood plasma samples of healthy volunteers who underwent a comprehensive medical examination and were found eligible for space flights was performed. As a result of directed mass spectrometric analysis, signals for 128 proteins, which accounted for nearly 40% of the total number of chromosome 13 gene products, were detected. The analysis of interindividual variation of concentrations of chromosome 13 proteins showed the presence of a pool comprising 41 proteins with a low variation (CV < 30%), which can potentially be used as biomarkers. 相似文献
8.
Hiroaki Kataoka Kazuki Nabeshima Naoto Komada Masashi Koono 《Virchows Archiv. B, Cell pathology including molecular pathology》1989,57(1):157-165
Two new human cell lines, RCM-1 and CoCM-1, have been established from primary colorectal adenocarcinomas. Both cell lines
were unique in that the cultures secreted trypsin inhibitors in vitro. The activities of these inhibitors were accumulated
in serum-free media of both cell lines over a period of several days. Two inhibitors (PI-1 and PI-2) were isolated from serum-free
conditioned medium in which RCM-1 was grown by anion-exchange and gel filtration high-performance liquid chromatography. PI-1
inhibited trypsin and chymotrypsin strongly, and pancreatic elastase weakly. Its molecular weight was about 57 kilodaltons
(Kd) as determined by gel filtration chromatography. It cross-reacted with the antiserum elicited against human α1-antitrypsin in double immunodiffusion. PI-1 corresponding to α1
- antitrypsin was also demonstrated immunohistochemically in both cell lines. PI-2 inhibited trypsin strongly, and chymotrypsin,
kallikrein and plasmin weakly. It had higher molecular weight (200–300 Kd) than that of PI-1, and did not crossreact with
antisera against human α1-antitrypsin, α2-macroglobulin, α1-antichymotrypsin, α2-plasmin inhibitor, inter-α-trypsin inhibitor and urinary trypsin inhibitor. RCM-1 and CoCM-1 are the first colorectal adenocarcinoma
cell lines that secrete functionally active trypsin inhibitors, including α1-antitrypsin in vitro, and are useful for the study of tumor-cell derived proteinase inhibitors. 相似文献
9.
Elaković I Perisić T Canković-Kadijević M Matić G 《Cell biochemistry and function》2007,25(4):427-431
Correlation between the glucocorticoid receptor (GR) number and affinity for the ligand, as well as the relationship between these equilibrium binding parameters and body mass index, blood pressure, and age were examined in peripheral blood mononuclear cells (PBMC) of healthy human subjects. It was found that the only statistically significant correlation was that between the GR number per cell and equilibrium dissociation constant, K(d) (r = 0.84, p < 0.0001). This observation implies the existence of a compensatory mechanism providing for lower GR affinity in individuals that have more receptor sites in circulating mononuclear cells and vice versa. This compensatory phenomenon together with considerable interindividual variation (GR number per cell ranging from 1391 to 15133, CV = 58.62%; and K(d) from 2.5 to 98.6 nM, CV = 80.87%), reflects plasticity of the glucocorticoid system. The results pose the question of whether this compensatory mechanism observed in healthy human subjects persists in pathophysiological states associated with glucocorticoid hormone actions and suggest that tissue sensitivity to glucocorticoids could be better predicted by the sign and magnitude of the correlation between the two receptor equilibrium binding parameters than by each of them separately. 相似文献
10.
《Expert review of proteomics》2013,10(1):139-145
The dramatic progress in mass spectrometry-based methods of protein identification has triggered a new quest for disease-associated biomarkers. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and its variant surface-enhanced laser desorption/ionization mass spectrometry, provide effective means to explore the less studied information slice of the human serum proteome – low-molecular-weight proteins and peptides. These low-molecular-weight proteins and peptides are promising for the detection of important biomarkers. Due to the significant experimental problems imposed by high-abundance and high-molecular-weight proteins, it is important to effectively remove these species prior to mass spectrometry analysis of the low-molecular-weight serum and plasma proteomes. In this review, the advantages afforded by recently introduced methods for prefractionation of serum, as they pertain to the detection and identification of biomarkers, will be discussed. 相似文献
11.
Chertov O Simpson JT Biragyn A Conrads TP Veenstra TD Fisher RJ 《Expert review of proteomics》2005,2(1):139-145
The dramatic progress in mass spectrometry-based methods of protein identification has triggered a new quest for disease-associated biomarkers. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and its variant surface-enhanced laser desorption/ionization mass spectrometry, provide effective means to explore the less studied information slice of the human serum proteome -- low-molecular-weight proteins and peptides. These low-molecular-weight proteins and peptides are promising for the detection of important biomarkers. Due to the significant experimental problems imposed by high-abundance and high-molecular-weight proteins, it is important to effectively remove these species prior to mass spectrometry analysis of the low-molecular-weight serum and plasma proteomes. In this review, the advantages afforded by recently introduced methods for prefractionation of serum, as they pertain to the detection and identification of biomarkers, will be discussed. 相似文献
12.
A rabbit antibody against the light-chain of guinea-pig high-molecular-weight (HMW) kininogen, which was specific to HWM kininogen and did not recognize low-molecular-weight kininogen, was prepared. This antibody demonstrated the presence of HMW kininogen antigen at the interstitial-tissue space in the guinea-pig skin by means of immunohistochemistry. The interstitial-tissue HMW kininogen antigen was extracted from the skin. This antigen molecule in the skin extract behaved identically as HWM kininogen of plasma in slab-polyacrylamide gel electrophoresis under the presence of sodium dodecyl sulfate followed by immunoblotting. Therefore, it was concluded that HMW kininogen was present in the interstitial-tissue fluid in the skin. The amount of HMW kininogen in the skin extract was quantified by a sandwich enzyme-linked immunosorbent assay with the anti-light-chain antibody and a goat anti-guinea-pig HMW kininogen antibody. On the assumption that the interstitial-tissue volume is 50 ml/100 g wet skin tissue, the average concentration of HMW kininogen in the interstitial-tissue fluid of the skin was calculated to be 23% of the plasma concentration. On the other hand, the proportion of intravascular HMW kininogen (derived from blood remaining in the vessels of the harvested skin) in relation to the total HMW kininogen in the skin extract was quantified by measuring the radio-labelled HMW kininogen which had been injected intravenously as a tracer of the intravascular HMW kininogen. About 5% of the total HMW kininogen in the skin extract was calculated to be derived from the intravascular blood volume of the skin, indicating that the majority of the HMW kininogen in the skin extract was derived from the extravascular-tissue space. 相似文献
13.
Urine protein profile of IgA nephropathy patients may predict the response to ACE-inhibitor therapy 总被引:1,自引:0,他引:1
Rocchetti MT Centra M Papale M Bortone G Palermo C Centonze D Ranieri E Di Paolo S Gesualdo L 《Proteomics》2008,8(1):206-216
This study was aimed at the search of urinary biomarkers which might help to predict the clinical response of IgA nephropathy (IgAN) patients to angiotensin converting enzyme inhibitors (ACEi). First, we studied the urinary proteome of 18 IgAN patients (toward 20 healthy controls) who had been chronically treated with ACEi by using 2-D PAGE coupled to nano-HPLC-ESI-MS/MS analysis. We identified 3 proteins, kininogen (p = 0.02), inter-alpha-trypsin-inhibitor heavy chain 4 (35 kDa fragment) (p = 0.02) and transthyretin (p<0.0001), whose urinary excretion was different in IgAN patients' responders when compared to those who had not responded to ACEi. A reduction of daily proteinuria >50% and a stable renal function over time were used to classify patients as responders. Then, we adopted immunoblotting to confirm the predictive power of one of the above proteins, kininogen, in 20 patients with biopsy-proven IgAN, before starting any therapy. Thus, we confirmed that very low levels of kininogen urine excretion were indeed predictive of an inadequate or absent clinical response to ACEi therapy of IgAN patients, after 6-month follow-up. Concluding, the analysis of urine proteome of IgAN patients generated a set of proteins which distinguished subjects responsive to ACEi from those unresponsive to the inhibition of renin-angiotensin system (RAS). 相似文献
14.
Peripheral blood mononuclear cells (PBMCs) are main actors in inflammatory processes and linked to many diseases, including rheumatoid arthritis, atherosclerosis, asthma, HIV and cancer. Moreover, they seem an interesting ‘surrogate tissue’ that can be used in biomarker discovery. In order to get a good experimental design for quantitative expression studies, the knowledge of the interindividual variation is an essential part. Therefore, PBMCs were isolated from 24 healthy volunteers (15 males, 9 females, ages 63–86) with no clinical signs of inflammation. The extracted proteins were separated using the two dimensional difference in gel electrophoresis technology (2D-DIGE), and the gel images were processed with the DeCyder 2D software. Protein spots present in at least 22 out of 24 healthy volunteers were selected for further statistical analysis. Determination of the coefficient of variation (CV) of the normalized spot volume values of these proteins, reveals that the total variation of the PBMC proteome varies between 12,99% to 148,45%, with a mean value of 28%. A supplemental look at the causes of technical variation showed that the isolation of PBMCs from whole blood is the factor which influences the experimental variance the most. This isolation should be handled with extra care and an additional washing step would be beneficial. Knowing the extent of variation, we show that at least 10 independent samples per group are needed to obtain statistical powerful data. This study demonstrates the importance of considering variance of a human population for a good experimental design for future protein profiling or biomarker studies. 相似文献
15.
Zhang X Guo Y Song Y Sun W Yu C Zhao X Wang H Jiang H Li Y Qian X Jiang Y He F 《Proteomics》2006,6(19):5260-5268
Normal Chinese Liver Proteome Expression Profile is one of the major parts of Human Liver Proteome Project. Before starting the studies, it is necessary to examine the interindividual variation of normal liver proteome and evaluate the minimal size of samples for proteomic analysis. In this study, normal liver samples from ten individual volunteers were collected and the proteome profiles of these samples were analyzed using 2-D difference gel electrophoresis (DIGE) combined with MALDI-TOF/TOF MS. The individual liver tissue lysates were labeled with Cy3 and Cy5 while the pooled sample was labeled with Cy2 as an internal standard, which minimized gel-to-gel variation. After analysis by the DeCyder software, up to 2056 protein spots were detected on the master gel. The CV of standardized abundance was calculated for the protein spots that were matched across all ten gels. The CV values of these protein spots ranged from 6.4 to 108.5% and the median CV was approximately 19%, which demonstrated that the protein expression of normal liver among different individuals was relatively stable. The eight proteins with CV values over 50% were identified which would be a caveat when considering these proteins as potential disease-related markers. Moreover, the one-way ANOVA feature showed a correlation between sample size and individual variations. The results showed that when the sample size exceeded 7, the individual variations were not significant to the whole pool. Our results are an important basis for liver protein expression profiles and comparative proteomics of liver disease. 相似文献
16.
R. Štěpánková J. Šinkora T. Hudcovic H. Kozáková H. Tlaskalová-Hogenová 《Folia microbiologica》1998,43(5):531-534
The aim of the study was to compare the phenotype of lymphocyte subpopulations of the GALT (gut-associated lymphatic tissue)
in germfree (GF) and conventionally (CV) reared rats,i.e. to analyze the effect of microbial colonization on the development of intestinal lymphocyte subsets. Surface marker characteristics
were studied in cell suspensions isolated from Peyer’s patches, mesenteric lymph nodes, spleen and the intraepithelial lymphocyte
compartment of 2- and 12-month old inbred AVN rats. The pattern of T lymphocyte phenotypes in Peyer’s patches, mesenteric
lymph nodes and spleen determined by FACS analysis did not reveal differences between GF and CV rats. In contrast, a 2-month
conventionalization of GF rats led to substantial changes in the composition of intestinal intraepithelial lymphocyte subsets
(IELs): increase of CD4+, CD8α+, CD8β+, TcR α/β+ bearing lymphocytes was observed after colonization of rats with normal microflora. Surprisingly, the relative numbers of
lymphocytes bearing TcR γ/δ+ did not change during conventionalization. The effect of aging was also studied and differences in IELs composition of aged
(GF) and (CV) rats were found to be more pronounced: 6,6% and 30% of lymphocytes bearing TcR α/β were present among IELs in
two-month old GF and CV rats, respectively. 30% of IELs in 2-month old GF rats, 80% of IEL from 12-month old CV rats were
found to bear TcR α/β. This finding demonstrates that during conventionalization and aging the TcR α/β bearing population
of IELs substantially expands. It suggests that mainly this lymphocyte subset responds to microflora stimuli and is probably
involved in the protection of the epithelial cell layer of intestinal mucosa. 相似文献
17.
18.
Eivind Larssen Cato Brede Anne Bjørnstad Hjelle Kjell Birger Øysæd Anne Bolette Tjensvoll Roald Omdal Peter Ruoff 《Proteomics》2015,15(1):10-15
The cerebrospinal fluid (CSF) proteome is of great interest for investigation of diseases and conditions involving the CNS. However, the presence of high‐abundance proteins (HAPs) can interfere with the detection of low‐abundance proteins, potentially hindering the discovery of new biomarkers. Therefore, an assessment of the CSF subproteome composition requires depletion strategies. Existing methods are time consuming, often involving multistep protocols. Here, we present a rapid, accurate, and reproducible method for preparing the CSF proteome, which allows the identification of a high number of proteins. This method involves acetonitrile (ACN) precipitation for depleting HAPs, followed by immediate trypsination. As an example, we demonstrate that this method allows discrimination between multiple sclerosis patients and healthy subjects. 相似文献
19.
Renal cortical high-molecular-weight renin (Mw:60,000) of the dog is a complex of renin (low-molecular-weight renin; Mw:40,000) and a renin binding protein. We detected an enzyme-like substance that catalyzes the conversion from high- into low-molecular-weight renin. When the renal cortical extract was added to the high-molecular-weight renin and the preparation incubated at 37 degrees C for 30 min, the high-molecular-weight renin was converted into the low-molecular-weight form. No such conversion occurred in the case of renal medullary extract. This converting substance was fractionated using concanavalin A Sepharose, 70% ammonium sulfate saturation and DEAE-cellulose chromatography. The converting activity was inhibited by potassium tetrathionate, N-ethylmaleimide and 5,5'-dithiobis-(2-nitrobenzoic acid). These events suggest that this substance is an enzyme possessing sulfhydryl moieties. However, a cathepsin B inhibitor leupeptin did not affect the activity. Accordingly, the high-molecular-weight renin converting enzyme, which is sensitive to sulfhydryl oxidation, may explain the mechanism of interconversion between high- and low-molecular-weight renin involving the oxidation-reduction of tissue sulfhydryl groups. 相似文献
20.
Accelerating effect of zinc ions on the surface-mediated activation of factor XII and prekallikrein 总被引:3,自引:0,他引:3
The effect of zinc ions on the surface-mediated activation of factor XII and prekallikrein was studied, using the contact system reconstituted with the purified proteins from bovine and human plasmas. The sulfatide-mediated activation of factor XII and prekallikrein in the presence of high-molecular-weight (HMW) kininogen was remarkably accelerated by 10(-5) M zinc ions. This accelerating effect was observed only in the presence of HMW kininogen. The kinetic analysis of the accelerating effect of zinc ions demonstrated that zinc ions reduce the Km values and increase the Vmax values on the activation of factor XII by kallikrein and on the activation of prekallikrein by factor XIIa. The value of Vmax/Km increased 26.4-fold in the former reaction and 2.8-fold in the latter reaction, indicating that zinc ions accelerate mainly the activation of factor XII by kallikrein. In the presence of 5 x 10(-4) M zinc ions, typical difference spectra due to a red shift of tryptophan and/or tyrosine residues were observed for HMW kininogen and its derivatives but not low-molecular-weight (LMW) kininogen. Since the concentration of zinc ions required to induce the difference spectra is comparable with that to enhance the activation of factor XII and prekallikrein, it appears that there is some correlation between the conformational change of HMW kininogen and the enhancement of the activation. 相似文献