Pub Date : 2008-06-01DOI: 10.2198/JELECTROPH.52.39
N. Okayama, Y. Suehiro, Koji Ueno, S. Hazama, M. Oka, Y. Hinoda
It has been shown that a non-synonymous polymorphism Asp299Gly of Toll-like receptor-4 (TLR-4) gene could be associated with lipopolysaccharide hyporesponsiveness and decreased risk of atherosclerosis in Caucasians. However, this polymorphism is not detected in Asians including Japanese and hence considered to be a population-specific one. To investigate if there are common non-synonymous SNPs of TLR-4 gene in Japanese, 100 DNA samples randomly selected from 613 healthy volunteers were sequenced for exons 2 and 3. As a result, Thr175Ala (A690G) was found in 6 out of 100 samples. Genotyping of Thr175Ala was then performed for 842 DNA samples from 613 healthy volunteers and 229 patients with ulcerative colitis. Frequency of heterozygote (Thr/Ala) or homozygote (Ala/Ala) in healthy subjects was 2.3% (14/613) or 0% (0/613), respectively. There was no significant difference of the genotype distribution between ulcerative colitis patients and age- and gender-matched controls. Furthermore, Thr175 is known to be involved in a potential N-linked glycosylation site and hence the polymorphism might affect the function of TLR-4. These data suggest that Thr175Ala is a common non-synonymous polymorphism of TLR-4 gene in a Japanese population.
{"title":"Thr175Ala of Toll-like receptor-4 gene is a common non-synonymous single nucleotide polymorphism in a Japanese population","authors":"N. Okayama, Y. Suehiro, Koji Ueno, S. Hazama, M. Oka, Y. Hinoda","doi":"10.2198/JELECTROPH.52.39","DOIUrl":"https://doi.org/10.2198/JELECTROPH.52.39","url":null,"abstract":"It has been shown that a non-synonymous polymorphism Asp299Gly of Toll-like receptor-4 (TLR-4) gene could be associated with lipopolysaccharide hyporesponsiveness and decreased risk of atherosclerosis in Caucasians. However, this polymorphism is not detected in Asians including Japanese and hence considered to be a population-specific one. To investigate if there are common non-synonymous SNPs of TLR-4 gene in Japanese, 100 DNA samples randomly selected from 613 healthy volunteers were sequenced for exons 2 and 3. As a result, Thr175Ala (A690G) was found in 6 out of 100 samples. Genotyping of Thr175Ala was then performed for 842 DNA samples from 613 healthy volunteers and 229 patients with ulcerative colitis. Frequency of heterozygote (Thr/Ala) or homozygote (Ala/Ala) in healthy subjects was 2.3% (14/613) or 0% (0/613), respectively. There was no significant difference of the genotype distribution between ulcerative colitis patients and age- and gender-matched controls. Furthermore, Thr175 is known to be involved in a potential N-linked glycosylation site and hence the polymorphism might affect the function of TLR-4. These data suggest that Thr175Ala is a common non-synonymous polymorphism of TLR-4 gene in a Japanese population.","PeriodicalId":15059,"journal":{"name":"Journal of capillary electrophoresis","volume":"1 1","pages":"39-42"},"PeriodicalIF":0.0,"publicationDate":"2008-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"90926975","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2008-03-01DOI: 10.2198/JELECTROPH.52.19
H. Okusa, Y. Kodera, M. Oh‐ishi, Yutaka Minamida, M. Tsuchida, N. Kavoussi, Kazumasa Matsumoto, Takefumi Sato, M. Iwamura, T. Maeda, S. Baba
Urine cytology is the gold standard for detection of bladder cancer. However, it has poor sensitivity, and currently several diagnostic markers have high false-positive rates. One of the problems with bladder cancer is high locoregional recurrence after surgery. Therefore new diagnostic markers of bladder cancer are needed. We have conducted a comprehensive study of high molecular mass (HMM) protein expression in bladder cancer tissue by means of agarose two-dimensional gel electrophoresis (agarose 2-DE) followed by the analysis of liquid chromatography tandem mass spectroscopy. We successfully identified 53 proteins from cancer tissue and 33 proteins from normal mucosa in the HMM area of the 2DE gel, and they were classified as proteins of transcription, translation or the cytoskeleton. Among them, eight differentially regulated proteins were selected as new marker candidates, and we performed experiments to validate their use as diagnostic markers. The results of Western blotting showed that these proteins except for α spectrin were significantly upregulated in bladder cancer tissue, and one of them, structural maintenance of chromosomes protein 3 (SMC3), was specifically detected in urine samples from bladder cancer patients. On the other hand, the results of Immunohistochemistry showed that expression of neuroblast differentiation-associated protein AHNAK (AHNAK) was significantly increased and that of plectin-1 was significantly decreased in the bladder cancer specimens compared with normal bladder mucosa specimens. However, no significant difference was observed in other proteins. These results suggest that AHNAK and Plectin-1 would be the potential markers for urine cytology and SMC3 is possible to be that for diagnosis marker for urinalysis.
{"title":"Searching for new biomarkers of bladder cancer based on proteomic analysis","authors":"H. Okusa, Y. Kodera, M. Oh‐ishi, Yutaka Minamida, M. Tsuchida, N. Kavoussi, Kazumasa Matsumoto, Takefumi Sato, M. Iwamura, T. Maeda, S. Baba","doi":"10.2198/JELECTROPH.52.19","DOIUrl":"https://doi.org/10.2198/JELECTROPH.52.19","url":null,"abstract":"Urine cytology is the gold standard for detection of bladder cancer. However, it has poor sensitivity, and currently several diagnostic markers have high false-positive rates. One of the problems with bladder cancer is high locoregional recurrence after surgery. Therefore new diagnostic markers of bladder cancer are needed. We have conducted a comprehensive study of high molecular mass (HMM) protein expression in bladder cancer tissue by means of agarose two-dimensional gel electrophoresis (agarose 2-DE) followed by the analysis of liquid chromatography tandem mass spectroscopy. We successfully identified 53 proteins from cancer tissue and 33 proteins from normal mucosa in the HMM area of the 2DE gel, and they were classified as proteins of transcription, translation or the cytoskeleton. Among them, eight differentially regulated proteins were selected as new marker candidates, and we performed experiments to validate their use as diagnostic markers. The results of Western blotting showed that these proteins except for α spectrin were significantly upregulated in bladder cancer tissue, and one of them, structural maintenance of chromosomes protein 3 (SMC3), was specifically detected in urine samples from bladder cancer patients. On the other hand, the results of Immunohistochemistry showed that expression of neuroblast differentiation-associated protein AHNAK (AHNAK) was significantly increased and that of plectin-1 was significantly decreased in the bladder cancer specimens compared with normal bladder mucosa specimens. However, no significant difference was observed in other proteins. These results suggest that AHNAK and Plectin-1 would be the potential markers for urine cytology and SMC3 is possible to be that for diagnosis marker for urinalysis.","PeriodicalId":15059,"journal":{"name":"Journal of capillary electrophoresis","volume":"35 1","pages":"19-24"},"PeriodicalIF":0.0,"publicationDate":"2008-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"87918779","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2008-03-01DOI: 10.2198/JELECTROPH.52.25
Yuji Hirano, A. Kobayashi, Akira Haruki, H. Haga, M. Hara, Toyoji Sato, T. Ōhashi, K. Sasagawa, T. Miura, Yukio Shima, S. Watanabe
We studied serum samples which showed alkaline phosphatase (ALP) activity at or near the origin of the polyacrylamide gel electrophoresis. 1) Sucrose monolaurate (final concentration, 1.7%) treatment produced a new ALP band (SM-ALP band). 2) The SM-ALP band disappeared from the original region with anti-human placental ALP antibody and appeared at the high molecular region. 3) ALP activity of SM-ALP band was completely inactivated by heat treatment (at 65°C for 10 min). 4) The molecular mass was estimated to be approximately 350 kDa by a 5-20% (linear gradient) polyacrylamide slab gel electrophoresis. We concluded that the SM-ALP band was intestinal ALP tetramer.
{"title":"Characterization of serum alkaline phosphatase separated at or near the origin in polyacrylamide gel using detergents, antibodies and neuraminidase","authors":"Yuji Hirano, A. Kobayashi, Akira Haruki, H. Haga, M. Hara, Toyoji Sato, T. Ōhashi, K. Sasagawa, T. Miura, Yukio Shima, S. Watanabe","doi":"10.2198/JELECTROPH.52.25","DOIUrl":"https://doi.org/10.2198/JELECTROPH.52.25","url":null,"abstract":"We studied serum samples which showed alkaline phosphatase (ALP) activity at or near the origin of the polyacrylamide gel electrophoresis. 1) Sucrose monolaurate (final concentration, 1.7%) treatment produced a new ALP band (SM-ALP band). 2) The SM-ALP band disappeared from the original region with anti-human placental ALP antibody and appeared at the high molecular region. 3) ALP activity of SM-ALP band was completely inactivated by heat treatment (at 65°C for 10 min). 4) The molecular mass was estimated to be approximately 350 kDa by a 5-20% (linear gradient) polyacrylamide slab gel electrophoresis. We concluded that the SM-ALP band was intestinal ALP tetramer.","PeriodicalId":15059,"journal":{"name":"Journal of capillary electrophoresis","volume":"90 1","pages":"25-27"},"PeriodicalIF":0.0,"publicationDate":"2008-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"83913136","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Dielectrophoresis (DEP) is the motion of a matter caused by polarization effects in a non-uniform electric field. Recently, studies on DEP are promoted in various medical fields, including separation and characterization of biological cells. It is very important but not easy to measure this motive force, so-called dielectrophoretic force (DEP force). In general, the DEP force generated by the designed electrodes is analyzed by the computer simulation that employs the finite element analysis method. However, it does not always calculate the correct DEP force. Therefore, we propose the method of measuring the DEP force accurately based on the null method. The experiment is conducted with the dielectrophoretic device (DEP device) in which microfabricated electrodes were formed and which contains the polystyrene particles exhibiting the negative DEP in distilled water in a non-uniform electric field. The displacement of the particle reaches a steady state 15 min. after the change of applied voltage v, angle θ or frequency f. The equilibrium state of the particle in a non-uniform electric field can be reached at any place by adjusting both of angles θ and φ, where θ is the “Pitch” angle and φ the “Yaw” angle for the device. The proposed method can measure the incredibly minute DEP force ranging from 25 fN (femto-Newton) at θ=5° to 298 fN at θ=90°, the accuracy of which is determined by the static friction and rolling friction between particles and the inner floor of the DEP device.
{"title":"Measurement of dielectrophoretic force by employing controllable gravitational force","authors":"H. Imasato, T. Yamakawa","doi":"10.2198/JELECTROPH.52.1","DOIUrl":"https://doi.org/10.2198/JELECTROPH.52.1","url":null,"abstract":"Dielectrophoresis (DEP) is the motion of a matter caused by polarization effects in a non-uniform electric field. Recently, studies on DEP are promoted in various medical fields, including separation and characterization of biological cells. It is very important but not easy to measure this motive force, so-called dielectrophoretic force (DEP force). In general, the DEP force generated by the designed electrodes is analyzed by the computer simulation that employs the finite element analysis method. However, it does not always calculate the correct DEP force. Therefore, we propose the method of measuring the DEP force accurately based on the null method. The experiment is conducted with the dielectrophoretic device (DEP device) in which microfabricated electrodes were formed and which contains the polystyrene particles exhibiting the negative DEP in distilled water in a non-uniform electric field. The displacement of the particle reaches a steady state 15 min. after the change of applied voltage v, angle θ or frequency f. The equilibrium state of the particle in a non-uniform electric field can be reached at any place by adjusting both of angles θ and φ, where θ is the “Pitch” angle and φ the “Yaw” angle for the device. The proposed method can measure the incredibly minute DEP force ranging from 25 fN (femto-Newton) at θ=5° to 298 fN at θ=90°, the accuracy of which is determined by the static friction and rolling friction between particles and the inner floor of the DEP device.","PeriodicalId":15059,"journal":{"name":"Journal of capillary electrophoresis","volume":"12 1","pages":"1-8"},"PeriodicalIF":0.0,"publicationDate":"2008-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"78256984","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2008-03-01DOI: 10.2198/JELECTROPH.52.29
H. Kajiwara, Masatoshi Nakamura
Proteins from silkworm (Bombyx mori L.) silk glands, fat bodies, and hemolymph were analyzed by matrix assisted laser desorption ionization time-of-flight mass spectrometry using sinapinic acid as a matrix. Peak percentages were calculated for each peak from m/z 2,000 to m/z 20,000 against the sum of all observed peak intensities; hierarchical cluster analysis and heat map analysis were performed using R-statistics software. For the silk glands, clusters obtained from MALDI-TOF-MS analysis were consistented with the morphological classification. Hierarchical clustering using MALDI-TOF-MS data was considered applicable to the small and sequential biopsies. In hemolymph, obvious clusters were observed from the anterior part to posterior part, and the proteins in hemolymph were not considered homogeneous throughout the body. While the clusters obtained from fat body analysis were weaker than those of silk glands and hemolymph, three clusters were observed. Based on the protein profile, fat bodies were also considered as differentiated tissue.
{"title":"Hierarchical cluster analyses and heat map analyses of silkworm tissues using R-statistics","authors":"H. Kajiwara, Masatoshi Nakamura","doi":"10.2198/JELECTROPH.52.29","DOIUrl":"https://doi.org/10.2198/JELECTROPH.52.29","url":null,"abstract":"Proteins from silkworm (Bombyx mori L.) silk glands, fat bodies, and hemolymph were analyzed by matrix assisted laser desorption ionization time-of-flight mass spectrometry using sinapinic acid as a matrix. Peak percentages were calculated for each peak from m/z 2,000 to m/z 20,000 against the sum of all observed peak intensities; hierarchical cluster analysis and heat map analysis were performed using R-statistics software. For the silk glands, clusters obtained from MALDI-TOF-MS analysis were consistented with the morphological classification. Hierarchical clustering using MALDI-TOF-MS data was considered applicable to the small and sequential biopsies. In hemolymph, obvious clusters were observed from the anterior part to posterior part, and the proteins in hemolymph were not considered homogeneous throughout the body. While the clusters obtained from fat body analysis were weaker than those of silk glands and hemolymph, three clusters were observed. Based on the protein profile, fat bodies were also considered as differentiated tissue.","PeriodicalId":15059,"journal":{"name":"Journal of capillary electrophoresis","volume":"22 1","pages":"29-38"},"PeriodicalIF":0.0,"publicationDate":"2008-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"83922476","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Akira Sawada, Takeshi Ueno, Y. Kawashima, Eri Haruta-Satoh, M. Oh‐ishi, Y. Kodera, T. Maeda
SUMMARY Oxidative stress is implicated in a broad variety of chronic and acute diseases, including Alzheimer’s disease, arteriosclerosis and diabetes. In order to study oxidative stress at the proteome level proteins were labeled with Cy2NHS (the N-hydroxyl succinimidyl ester derivative of a cyanine dye reactive with the ε -amino group of lysine residues), or with Cy3- and Cy5HZ (hydrazide derivatives of the dyes reactive with protein carbonyls) and analyzed by two-dimensional fluorescence difference gel electrophoresis (2D-DIGE). Two samples were used for testing the CyHZ labeling; one is low molecular mass marker proteins artificially oxidized by NaOCl and the other renal cortex extracts of 30-week-old diabetes model OLETF (Otsuka Long-Evans Tokushima Fatty) and control LETO (Long-Evans Tokushima Otsuka) rats. Spot volumes V ni and contrasts c ni of 2D-DIGE images were quantified following background subtraction, where the suffices n (=2, 3 or 5) and i (=1, 2, ...) represent the Cy-signal and the Cy2-detected spot identification numbers, respectively. The contrasts c 3 i and c 5 i were on average about 0.53 times as high as c 2 i . Tak-ing the contrast into account, a simple parameter q ni was proposed for judging the quality of spot images. The difference in-gel analysis (DIA) algorithm should be used in combina-tion with the quality parameters to identify statistical outliers as candidates for abnormally carbonylated proteins. Inspection of the background flatness around the candidate spots is also required before making a conjecture as to whether the candidate protein is abnormally oxidized.
{"title":"Protein carbonyl detection by two-dimensional fluorescence difference gel electrophoresis","authors":"Akira Sawada, Takeshi Ueno, Y. Kawashima, Eri Haruta-Satoh, M. Oh‐ishi, Y. Kodera, T. Maeda","doi":"10.2198/JELECTROPH.52.9","DOIUrl":"https://doi.org/10.2198/JELECTROPH.52.9","url":null,"abstract":"SUMMARY Oxidative stress is implicated in a broad variety of chronic and acute diseases, including Alzheimer’s disease, arteriosclerosis and diabetes. In order to study oxidative stress at the proteome level proteins were labeled with Cy2NHS (the N-hydroxyl succinimidyl ester derivative of a cyanine dye reactive with the ε -amino group of lysine residues), or with Cy3- and Cy5HZ (hydrazide derivatives of the dyes reactive with protein carbonyls) and analyzed by two-dimensional fluorescence difference gel electrophoresis (2D-DIGE). Two samples were used for testing the CyHZ labeling; one is low molecular mass marker proteins artificially oxidized by NaOCl and the other renal cortex extracts of 30-week-old diabetes model OLETF (Otsuka Long-Evans Tokushima Fatty) and control LETO (Long-Evans Tokushima Otsuka) rats. Spot volumes V ni and contrasts c ni of 2D-DIGE images were quantified following background subtraction, where the suffices n (=2, 3 or 5) and i (=1, 2, ...) represent the Cy-signal and the Cy2-detected spot identification numbers, respectively. The contrasts c 3 i and c 5 i were on average about 0.53 times as high as c 2 i . Tak-ing the contrast into account, a simple parameter q ni was proposed for judging the quality of spot images. The difference in-gel analysis (DIA) algorithm should be used in combina-tion with the quality parameters to identify statistical outliers as candidates for abnormally carbonylated proteins. Inspection of the background flatness around the candidate spots is also required before making a conjecture as to whether the candidate protein is abnormally oxidized.","PeriodicalId":15059,"journal":{"name":"Journal of capillary electrophoresis","volume":"104 1","pages":"9-17"},"PeriodicalIF":0.0,"publicationDate":"2008-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"77208018","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
MicroRNAs (miRNAs) are relatively short, 22nt～, and endogenous non-coding RNAs. It is known to act as a sequence specific regulator of post-transcriptional gene expression in many eukaryotes, most of the time with some sequence mismatches against target genes. Recently, a couple of miRNAs have been shown to be involved in oncogenic pathway, development, or cell differentiation. miRNAs are now getting attention to its diverse regulatory and catalytic functions.Although many of the miRNA sequences have been registered in the public databases and their target genes are predicted by computational analysis, the empirical verification has to be done. One of the difficulties to study the miRNA function will be that its structural and regulatory characteristics.In order to overcome this difficulty and utilize miRNA as an experimental tool, we investigated a system to detect, knockdown, and / or overexpress miRNAs. First, we adopted bicyclic nucleic acid, Locked Nucleic Acid (LNA), to improve affinity and single nucleotide discrimination. Next, we established the system to introduce synthetic oligos or RNAs expressed from lentiviral vectors to cells. A novel approach based on these principles on miRNA and its pathway analysis will be discussed.
{"title":"機能解析を目的としたmicroRNAの検出／発現／阻害システムの確立 ～It’s a small miRNA World～","authors":"隆之 水谷, 山田 佳世子","doi":"10.2198/SBK.51.211","DOIUrl":"https://doi.org/10.2198/SBK.51.211","url":null,"abstract":"MicroRNAs (miRNAs) are relatively short, 22nt～, and endogenous non-coding RNAs. It is known to act as a sequence specific regulator of post-transcriptional gene expression in many eukaryotes, most of the time with some sequence mismatches against target genes. Recently, a couple of miRNAs have been shown to be involved in oncogenic pathway, development, or cell differentiation. miRNAs are now getting attention to its diverse regulatory and catalytic functions.Although many of the miRNA sequences have been registered in the public databases and their target genes are predicted by computational analysis, the empirical verification has to be done. One of the difficulties to study the miRNA function will be that its structural and regulatory characteristics.In order to overcome this difficulty and utilize miRNA as an experimental tool, we investigated a system to detect, knockdown, and / or overexpress miRNAs. First, we adopted bicyclic nucleic acid, Locked Nucleic Acid (LNA), to improve affinity and single nucleotide discrimination. Next, we established the system to introduce synthetic oligos or RNAs expressed from lentiviral vectors to cells. A novel approach based on these principles on miRNA and its pathway analysis will be discussed.","PeriodicalId":15059,"journal":{"name":"Journal of capillary electrophoresis","volume":"45 1","pages":"211-214"},"PeriodicalIF":0.0,"publicationDate":"2007-09-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"77270879","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2007-09-01DOI: 10.2198/JELECTROPH.51.21
T. Toda, Megumi Nakamura, H. Morisawa, M. Hirota
The free-radical theory predicts that the oxidative stress accelerates the rate of aging and increases the onset of degenerative disorders in the elderly. Dopaminergic neurons are especially vulnerable to age-related neuronal disorders due to reactive oxygen species generated in the pathway of dopamine metabolism. Biochemical changes occurring in substantia nigra of Parkinson's disease patients suggest that the oxidative-stress-induced cell damages may be involved in the neurodegeneration. In our previous researches, we found that the dephosphorylation of elongation factor-2 and phosphorylation of nuclear lamin A/C might be neuronal cell specific response to oxidative stress. (Nakamura et al. BBA, 1763(9), 977-989, 2006) The dephosphorylation and phosphorylation of those proteins are significant biomarkers for analyzing the molecular mechanisms of the stress response, however, such a phosphoproteome analysis is thought to be inappropriate for clinical investigation of neurodegeneration if it was not detectable in cerebrospinal fluid or serum of patients. Thus, we proceeded to the 2D-DIGE analysis of secretome, proteome of secreted proteins, using the culture system in which oxidative stress was applied to human SH-SY5Y neuroblastoma cells. As the result of our secretome analysis, we identified ubiquitin-activating enzyme E1, ubiquitin-conjugating enzyme E2 N, ubiquitin C-terminal hydrorase-L1, 14-3-3 protein isoforms, Rab GDP dissociation inhibitor β, Rho GDP-dissociation inhibitor 1, peroxiredoxin-2, glutathione S-transferase P, α enolase, LDH B chain as oxidative-stress-reporting biomarker candidates.
{"title":"Proteomic identification of oxidative-stress-reporting biomarkers differentially secreted from human neuroblastoma SH-SY5Y cells","authors":"T. Toda, Megumi Nakamura, H. Morisawa, M. Hirota","doi":"10.2198/JELECTROPH.51.21","DOIUrl":"https://doi.org/10.2198/JELECTROPH.51.21","url":null,"abstract":"The free-radical theory predicts that the oxidative stress accelerates the rate of aging and increases the onset of degenerative disorders in the elderly. Dopaminergic neurons are especially vulnerable to age-related neuronal disorders due to reactive oxygen species generated in the pathway of dopamine metabolism. Biochemical changes occurring in substantia nigra of Parkinson's disease patients suggest that the oxidative-stress-induced cell damages may be involved in the neurodegeneration. In our previous researches, we found that the dephosphorylation of elongation factor-2 and phosphorylation of nuclear lamin A/C might be neuronal cell specific response to oxidative stress. (Nakamura et al. BBA, 1763(9), 977-989, 2006) The dephosphorylation and phosphorylation of those proteins are significant biomarkers for analyzing the molecular mechanisms of the stress response, however, such a phosphoproteome analysis is thought to be inappropriate for clinical investigation of neurodegeneration if it was not detectable in cerebrospinal fluid or serum of patients. Thus, we proceeded to the 2D-DIGE analysis of secretome, proteome of secreted proteins, using the culture system in which oxidative stress was applied to human SH-SY5Y neuroblastoma cells. As the result of our secretome analysis, we identified ubiquitin-activating enzyme E1, ubiquitin-conjugating enzyme E2 N, ubiquitin C-terminal hydrorase-L1, 14-3-3 protein isoforms, Rab GDP dissociation inhibitor β, Rho GDP-dissociation inhibitor 1, peroxiredoxin-2, glutathione S-transferase P, α enolase, LDH B chain as oxidative-stress-reporting biomarker candidates.","PeriodicalId":15059,"journal":{"name":"Journal of capillary electrophoresis","volume":"90 1","pages":"21-26"},"PeriodicalIF":0.0,"publicationDate":"2007-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"84586311","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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