Thioredoxin Domain-containing Protein Research Articles

Comparative Proteomic Analysis of Bone Marrow Plasma Cells By Using Mass Spectrometry Based Bottom up Proteomic Strategies

Aim : As all other cancer types, multiple myeloma is a complex disease, yet its molecular mechanism that is not fully understood and needs to be furtherly investigated. Besides, the molecular mechanism responsible from the transition of healthy cells into malignant ones is still under investigation. In such comparative studies, human myeloma cell lines are preferred as positive control, while tonsillar plasma cells are used as negative control. The aim of this study is to perform a comparative proteomic analysis regarding the progression of Multiple Myeloma (MM) by using protein profiles of plasma cells obtained from MGUS, SMM, symptomatic myeloma patients, tonsil plasma cells obtained from healthy individuals and human myeloma cells of cell line origin to elucidate the underlying molecular mechanism involved in myeloma.Material and Methods: Five groups were adopted in the experiment. Of these groups, MM patients were classified mainly into three groups according to marrow plasma cell content (PCC) validated by flow cytometry: group1: 0-9, group 2: 10-20 and group 3: >20 %. The fourth group consisted of human myeloma cell line (HMCL) RPMI 8226 and was designated as positive control, while tonsil plasma cells were used as negative control and specified as the fifth group. Marrow samples were collected from 30 patients newly diagnosed with Multiple Myeloma ( n: 28 symptomatic and n: 1 smoldering (SMM)) and MGUS (n: 1)).Tonsil plasma cells were isolated from healthy tonsil tissue treated with trypsin enzyme and were then confirmed by using Selection Kit microbead specific for EasySep Human CD138 marker. Protein profile maps of groups were obtained via 2D gel electrophoresis and comparative analysis and detection of up/down regulated protein spots was performed by using PDQuest 8.01 software. Proteins from differently expressed protein spots were then identified using Matrix-assisted laser desorption/ionization mass spectroscopy (MALDI) by Peptide Mass Fingerprinting (PMF) analysisResults: By using bottom up strategies, nine of the significantly expressed protein spots were identified with PMF analyses. The identified proteins are as follows:, Calreticulin (ERp60), Endoplasmin/ tumor rejection antigen (ERp99), MZB1(Marginal zone B and B1 cell specific protein/ pERp1), Actin cytoplasmic1 (ACTB), Thioredoxin domain-containing protein 5 (TXNDC5/ERp46), Protein disulfide isomerase (PDI) and HLA class I histocompatibility antigen (MHC class I antigen A*2). According to the results, calreticulin (which functions as chaperones and is responsible for Ca+2regulation and cell proliferation) showed a weak positive correlation with increase in patient PC content. The level was lowest in normal B cells. Endoplasmin/ tumor rejection antigen (which is assigned as chaperones and function in apoptotic process and Ca+2 regulation)declined by increase in PC content in patient samples. Level was lowest in HMCL. Marginal zone B and B1 cell specific protein (which functions as chaperones and is involved in apoptotic process, oxidoreductase activity, regulation of Ca+2, cell proliferation and activates B cell quantiity) correlated with PC percentage and was highest in HCML and lowest in normal B cells. Actin cytoplasmic1 protein (which functions in cell proliferation)synthesis showed a minimum decline in synthesis by increase in PC content .Level was highest in normal B cells. Thioredoxin domain-containing protein 5 which plays role in apoptotic process and shows oxireductase activity within MM/MGUS plasma cells also displayed a positive correlation but we were not able to detect a difference in HMCL and normal B cells which were synthesized less than the patient samples. Protein disulfide isomerase (which acts as chaperones and plays part in apoptotic process and shows oxireductase function) has a weak negative correlation with PC content and synthesis was lowest in HMCL. HLA class I histocompatibility antigen production did not correlate with PC content , it was higher in normal B cells compared to HMCL.Conclusion: In conclusion in this study we were able to confirm our previous findings which were presented at ASH 2013 by comparing the results with normal tonsil CD138+ plasma cells and a HMCL. Our study results suggest that the proteins which are involved in Calcium metabolism, chaperone binding and oxidative stress are also associated with low vs. high proliferation profile in myeloma. DisclosuresNo relevant conflicts of interest to declare.

Abstract 4960: ERp46 (thioredoxin domain-containing protein 5, TXND5) promotes prostate cancer growth in vitro and in vivo

Abstract We have recently demonstrated that endoplasmic reticulum protein ERp46, a member of the protein disulfide isomerase family of oxidoreductases, TXND5, is overexpressed in human metastatic renal cell carcinoma. The expression and function of ERp46 in prostate cancer has not been studied. Using both in vitro and in vivo approaches, we explore the suitability of ERp46 as a potential therapeutic target in prostate cancer. Tissue microarray containing normal prostate epithelium (n = 9) and prostate cancer specimens from 57 patients was stained for ERp46 and the staining intensity (H-score) was determined. Human prostate adenocarcinoma 22Rv1 cells were used to generate gain- and loss-of-function models by stable ERp46 shRNA knockdown and ERp46 overexpression, respectively. In vitro, the doubling time and PSA production were determined. In vivo, xenografts of each subclone were established in nude mice (n = 10/group) to determine the longitudinal tumor growth and serum PSA values. Gene expression profiling of RNA isolated from 22Rv1 xenografts was performed using human whole genome HT-12 V4 BeadChip array (Illumina). Our results demonstrated that human prostate carcinoma samples of Gleason scores ≥7 showed strong cytoplasmic ERp46 staining which was significantly increased compared to normal prostatic tissue (p = 0.02). ERp46 staining in prostate tumors of Gleason scores ≤6, however, was not different compared to normal prostate tissue. The stably transfected human prostate carcinoma 22Rv1 cells expressed 89% knockdown of ERp46 protein expression (shERp46) or a 4-fold increase in ERp46 protein expression (ERp46+) compared to the respective control cells. In vitro, shERp46 cells proliferated slower, whereas ERp46+ cells exhibited accelerated growth compared to corresponding control cells (p<0.05). Similarly, the tumor volume of subcutaneously growing shERp46 cells in nude mice led to significantly slower tumor growth (p<0.0005, ANOVA). Vice versa, tumors from ERp46+ cells were significantly larger than the tumor volume of shControl-cell injected mice (p = 0.02, ANOVA). Gene expression analysis confirmed the downregulation and upregulation of ERp46 in the corresponding xenografts and also showed several candidate genes, including NAAA, SH3BP4 and ID1 that were reciprocally up-and downregulated. In conclusion, this is the first report to suggest a role for ERp46 as an oncogenic protein and potential therapeutic target in prostate cancer given its expression profile in human prostate cancer samples and its effect on prostate cancer cell growth. Funding was provided by Prostate Cancer Canada and McMaster Surgical Associates (JHP and WCMD) Citation Format: Jehonathan H. Pinthus, Sarah N. Hopmans, Stephanie Federov, Wilhelmina C. Duivenvoorden. ERp46 (thioredoxin domain-containing protein 5, TXND5) promotes prostate cancer growth in vitro and in vivo. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4960. doi:10.1158/1538-7445.AM2015-4960

The role of TXNDC5 in castration-resistant prostate cancer-involvement of androgen receptor signaling pathway.

Castration-resistant prostate cancer (CRPC) continues to be a major clinical problem and the mechanisms behind it remain unclear. Thioredoxin domain-containing protein 5 (TXNDC5) is involved in protein folding and chaperone activity, and its overexpression has been reported in multiple malignancies. In the current study, we demonstrated that TXNDC5 is up-regulated following long-term androgen-deprivation treatment (ADT) and is highly overexpressed in CRPC tumors compared with hormone-naive prostate cancer (PCa) cases. Functionally, in vitro and in vivo studies demonstrated that TXNDC5 overexpression promotes the growth of both androgen-dependent and castration-resistant PCa xenografts. Mechanistically, TXNDC5 directly interacts with the AR protein to increase its stability and thus enhances its transcriptional activity. TXDNC5-mediated CRPC growth can be fully abolished by AR inhibition, suggesting TXDNC5 up-regulation as an escape pathway for aberrant AR re-activation and CRPC growth in the milieu of low androgen. Indeed, we found that TXNDC5 is increased by ADT-induced hypoxia through HIF-1α in an miR-200b-dependent manner. Overall, we defined an important role of TXNDC5 in CRPC and further investigations are needed to screen TXNDC5 antagonists as a novel therapeutic approaches to treat PCa patients with CRPC.

Differential protective effects of extra virgin olive oil and corn oil in liver injury: A proteomic study

Extra virgin olive oil (EVOO) presents benefits against chronic liver injury induced by hepatotoxins such as carbon tetrachloride (CCl4); however, the protective mechanisms remain unclear. In the present study, a two-dimensional gel based proteomic approach was constructed to explore the mechanisms. Rats are injected with CCl4 twice a week for 4 weeks to induce liver fibrosis, and were fed laboratory chow plus 20% (w/w) of either corn oil or EVOO over the entire experimental period. Histological staining, MDA assay and fibrogenesis marker gene analysis illustrate that the CCl4-treated animals fed EVOO have a lower fibrosis and lipid peroxidation level in the liver than the corn oil fed group. The proteomic study indicates that the protein expression of thioredoxin domain-containing protein 12, peroxiredoxin-1, thiosulphate sulphurtransferase, calcium-binding protein 1, Annexin A2 and heat shock cognate 71 kDa protein are higher in livers from EVOO-fed rats with the CCl4 treatment compared with those from rats fed with corn oil, whereas the expression of COQ9, cAMP-dependent protein kinase type I-alpha regulatory subunit, phenylalanine hydroxylase and glycerate kinase are lower. Our findings confirmed the benefits of EVOO against chronic liver injury, which may be attributable to the antioxidant effects, hepatocellular function regulation and hepatic metabolism modification effects of EVOO.

Preliminary Results Of A Mass Spectrometry Based Bottom Up Proteomic Approach On Bone Marrow Plasma Cells From Patients With Multiple Myeloma (MM)

AimThe molecular mechanisms responsible from evolution to malignant monoclonal plasmacytosis is still under investigation. The aim of this prospective study was to analyze the proteomics profile of plasma cells obtained from MGUS, SMM and symptomatic myeloma patients to be able to investigate the differences at protein level between patients with low vs high plasma cell content (PCC). Material and MethodsMarrow samples were collected from 30 patients newly diagnosed with Multiple Myeloma (n: 28 symptomatic and n: 1 smoldering (SMM)) and n:1 MGUS) at Ankara University Department of Hematology. Plasma cells were isolated by CD138+ selection before protein extraction. The patients were classified mainly in to three groups according to marrow PCC by flow cytometry: group1: 0-9, group 2: 10-20 and group 3: >20 %. The protein profiles of these three groups were constructed and compared via 2D gel electrophoresis by using PDQuest 8.01 analysis. Up/down regulated protein spots were identified with Matrix-assisted laser desorption/ionization mass spectroscopy(MALDI) by Peptide Mass Fingerprinting (PMF) analysis ResultsAll protein spots that were detected according to PCC with PDQuest analysis are as follows: 135 spots in group 1 142 spots in group 2 and 145 spots in group 3 Among these spots, 27 spots with significant expression density difference (at least 2-fold) were detected between group 1 and 2, 36 spots between group 1 and group 3 and 28 spots between group 2 and group 3. 36 of these protein spots were were used in peptide isolation by using trypsin. PMF analyses were carried out in MALDI-TOF mass spectrometer. From these spots, eight proteins were identified by using Mascot: Endoplasmin (ERp99), Calreticulin(ERp60), MZB1(Marginal zone B and B1 cell specific protein/pERp1), Actin cytoplasmic1(ACTB), Myeloblastin (Leukocyte proteinase 3), Thioredoxin domain-containing protein 5 (TXNDC5/ERp46), Apoptosis regulator B-cell lymphoma 2 (Bcl-2) and Peroxiredoxin-4 ( Table 1, Figure 1). The remaining 28 spots are under investigation. In group 3 the density of protein spots which contain Calreticulin, MZB1, Myeloblastin and TXCDN5 increased, and which contain Actin and Endoplasmin decreased significantly (Table 1). There was a negative corelation between the Peroxiredoxin expression level and the PCC which resulted in disappearance as the percentage of PCC increased. Moreover, not only Peroxiredoxin, but also BCL-2 protein was detected only in the group with PCC >20 %. To avoid the changes that can be attributable to PC counts equal amounts of protein were analyzed from each group. ConclusionUntil now there is only one published study utilizing proteomics in MM and reports 268 proteins (Chun Hua Lu et al, J Proteomics Informatics 2010). Ours is the first proteomics study comparing plasma cell proteins with PCC. The functional properties of these proteins are summarized (Table 2). High protein production and folding plus Ca changes in MM support our findings on chaperons and Ca binding protein changes. Out of these eight proteins only bcl-2, MZB1 and calreticulin were previously reported to be involved in the biology of MM. [Display omitted] Table 2Chromosomal location and functional properties of the eight proteins detected ( ref: www.nextprot.org)ProteinChromosomal LocationChaperoneApoptotic processOxidoreductaseRegulation of Ca(2+)Cell proliferationCalreticulin19p13.2+++Endoplasmin12q23.3+++MZB15q31.2+++++Actin cytoplasmic17p22.1+TXNDC56p24.3++Peroxiredoxin -4Xp22.11+Bcl-218q21.33+++Myeloblastin19p13.3+ Disclosures:Beksac:Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Honoraria, Membership on an entity's Board of Directors or advisory committees; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees.

Identification of proteins that mediate the pro-viral functions of the interferon stimulated gene 15 in hepatitis C virus replication

In previous studies we identified the interferon stimulated gene 15 (ISG15) as a pro-viral host factor in the pathogenesis of hepatitis C virus (HCV) infection. However, the functional link between ISG15 and the HCV replication cycle is not well understood. Aim of the present study was to functionally analyze the role of ISG15 and to identify possible HCV promoting effector molecules. Isg15 suppression was investigated in the murine subgenomic HCV replicon (MH1) transfected with Isg15-specific siRNA and in C57BL/6 mice intravenously injected with lipid nanoparticles (LNP)-formulated siRNA. Interestingly, the LNP-formulated siRNA led to hepatocyte-specific knockdown of Isg15 in vivo, which mediated a hypo-responsiveness to endogenous and exogenous interferon. A label free proteome analysis accompanied by western blot and quantitative RT-PCR techniques led to identification of five candidate proteins (Heterogeneous nuclear ribonucleoprotein A3 (HnrnpA3), Heterogeneous nuclear ribonucleoprotein K (HnrnpK), Hydroxymethylglutaryl-CoA synthase (Hmgcs1), Isocitrate dehydrogenase cytoplasmic (Idh1) and Thioredoxin domain-containing protein 5 (Txndc5)) that are either involved in lipid metabolism or belong to the family of Heterogeneous nuclear ribonucleoprotein (Hnrnp). All candidate proteins are likely to be associated with the HCV replication complex. Furthermore treatment with HnrnpK-specific siRNA directly suppressed HCV replication in vitro. Taken together these data suggest that targeting Isg15 may represent an attractive novel therapeutic option for the treatment of chronic HCV infection.

Proteomic analysis of S-nitrosylation induced by 1-methyl-4-phenylpyridinium (MPP+)

BackgroundNitric oxide (NO) mediates its function through the direct modification of various cellular targets. S-nitrosylation is a post-translational modification of cysteine residues by NO that regulates protein function. Recently, an imbalance of S-nitrosylation has also been linked to neurodegeneration through the impairment of pro-survival proteins by S-nitrosylation.ResultsIn the present study, we used two-dimensional gel electrophoresis in conjunction with the modified biotin switch assay for protein S-nitrosothiols using resin-assisted capture (SNO-RAC) to identify proteins that are S-nitrosylated more intensively in neuroblastoma cells treated with a mitochondrial complex I inhibitor, 1-methyl-4-phenylpyridinium (MPP+). We identified 14 proteins for which S-nitrosylation was upregulated and seven proteins for which it was downregulated in MPP+-treated neuroblastoma cells. Immunoblot analysis following SNO-RAC confirmed a large increase in the S-nitrosylation of esterase D (ESD), serine-threonine kinase receptor-associated protein (STRAP) and T-complex protein 1 subunit γ (TCP-1 γ) in MPP+-treated neuroblastoma cells, whereas S-nitrosylation of thioredoxin domain-containing protein 5 precursor (ERp46) was decreased.ConclusionsThese results suggest that S-nitrosylation resulting from mitochondrial dysfunction can compromise neuronal survival through altering multiple signal transduction pathways and might be a potential therapeutic target for neurodegenerative diseases.

Proteomics and gene expression analyses of squalene-supplemented mice identify microsomal thioredoxin domain-containing protein 5 changes associated with hepatic steatosis

Squalene is an abundant hydrocarbon present in virgin olive oil. Previous studies showed that its administration decreased atherosclerosis and steatosis in male apoE-knock-out mice. To study its effects on microsomal proteins, 1g/kg/day of squalene was administered to those mice. After 10 weeks, hepatic fat content was assessed and protein extracts of microsomal enriched fractions from control and squalene-treated animals were analyzed by 2D-DIGE. Spots exhibiting significant differences were identified by peptide fingerprinting and MSMS analysis. Squalene administration modified the expression of thirty-one proteins involved in different metabolic functions and increased the levels of those involved in vesicle transport, protein folding and redox status. Only mRNA levels of 9 genes (Arg1, Atp5b, Cat, Hyou1, Nipsnap1, Pcca, Pcx, Pyroxd2, and Txndc5) paralleled these findings. No such mRNA changes were observed in wild-type mice receiving squalene. Thioredoxin domain-containing protein 5 (TXNDC5) protein and mRNA levels were significantly associated with hepatic fat content in apoE-ko mice. These results suggest that squalene action may be executed through a complex regulation of microsomal proteins, both at the mRNA and post-transcriptional levels and the presence of apoE may change the outcome. Txndc5 reflects the anti-steatotic properties of squalene and the sensitivity to lipid accumulation.

Isolation and characterization of a thioredoxin domain-containing protein 12 from orange-spotted grouper, Epinephelus coioides

Thioredoxin domain-containing protein 12 (Txndc12) belongs to the thioredoxin superfamily, and has roles in redox regulation, defense against oxidative stress, refolding of disulfide-containing proteins, and regulation of transcription factors. In this study, a thioredoxin domain-containing protein 12 was cloned from the marine fish grouper, Epinephelus coioides by RACE PCR, named as Ec-Txndc12. The Ec-Txndc12 encodes 173 amino acid residues with signal peptide in its N-terminal and a thioredoxin (Trx) domain that is homologous with some genes in Mus musculus, Xenopus laveis, etc. Ec-Txndc12 mRNA is predominately expressed in liver, brain and muscle. The expression of Ec-Txndc12 was up-regulated in the liver of grouper challenged with SGIV. In order to elucidate its biological functions, Ec-Txndc12 was recombined and expressed in Escherichia coli BL21 (DE3). The rEc-Txndc12 fusion protein was demonstrated to possess the antioxidant activity. The grouper spleen (GS) cells were treated with a high concentration of rEc-Txndc12 (30 μg/ml), which significantly enhanced cells viability under oxidative damage caused by viral infection. These results together indicated that Ec-Txndc12 could function as an important antioxidant in a physiological context, and might be involved in the responses to viral challenge.

Differential proteome analysis of human embryonic kidney cell line (HEK-293) following mycophenolic acid treatment

BackgroundMycophenolic acid (MPA) is widely used as a post transplantation medicine to prevent acute organ rejection. In the present study we used proteomics approach to identify proteome alterations in human embryonic kidney cells (HEK-293) after treatment with therapeutic dose of MPA. Following 72 hours MPA treatment, total protein lysates were prepared, resolved by two dimensional gel electrophoresis and differentially expressed proteins were identified by QTOF-MS/MS analysis. Expressional regulations of selected proteins were further validated by real time PCR and Western blotting.ResultsThe proliferation assay demonstrated that therapeutic MPA concentration causes a dose dependent inhibition of HEK-293 cell proliferation. A significant apoptosis was observed after MPA treatment, as revealed by caspase 3 activity. Proteome analysis showed a total of 12 protein spots exhibiting differential expression after incubation with MPA, of which 7 proteins (complement component 1 Q subcomponent-binding protein, electron transfer flavoprotein subunit beta, cytochrome b-c1 complex subunit, peroxiredoxin 1, thioredoxin domain-containing protein 12, myosin regulatory light chain 2, and profilin 1) showed significant increase in their expression. The expression of 5 proteins (protein SET, stathmin, 40S ribosomal protein S12, histone H2B type 1 A, and histone H2B type 1-C/E/F/G/I) were down-regulated. MPA mainly altered the proteins associated with the cytoskeleton (26%), chromatin structure/dynamics (17%) and energy production/conversion (17%). Both real time PCR and Western blotting confirmed the regulation of myosin regulatory light chain 2 and peroxiredoxin 1 by MPA treatment. Furthermore, HT-29 cells treated with MPA and total kidney cell lysate from MMF treated rats showed similar increased expression of myosin regulatory light chain 2.ConclusionThe emerging use of MPA in diverse pathophysiological conditions demands in-depth studies to understand molecular basis of its therapeutic response. The present study identifies the myosin regulatory light chain 2 and peroxiredoxin 1 along with 10 other proteins showing significant regulation by MPA. Further characterization of these proteins may help to understand the diverse cellular effects of MPA in addition to its immunosuppressive activity.

Proteomic analysis reveals molecular biological details in varioliform gastritis withoutHelicobacter pyloriinfection

To investigate and elucidate the molecular mechanism underlying varioliform gastritis for early detection, prevention and intervention of gastric cancer. A combination of two-dimensional gel electrophoresis and mass spectrometry was used to detect the differentially expressed proteins between varioliform gastritis and matched normal mucosa. The selected proteins were confirmed by Western blotting and reverse transcription polymerase chain reaction (RT-PCR) in additional samples and the function of some proteins in varioliform gastritis was analyzed by bio-method preliminarily. We identified 21 differentially expressed proteins in varioliform gastritis, and compared them with matched normal mucosa. Eleven proteins were upregulated and ten downregulated in varioliform gastritis when compared with the same proteins in individual-matched normal gastric mucosa. These proteins are related to metabolism, oxidation, cytoskeleton, apoptosis, signal transduction and other aspects of cells. Two novel proteins, thioredoxin domain-containing protein 5 (TXNDC5) upregulated in varioliform gastritis, and neuropolypeptide h3 [phosphatidylethanolamine-binding protein 1 (PEBP1)] downregulated in varioliform gastritis, were further investigated. Their expressions were validated by Western blotting and RT-PCR in 12 cases of varioliform gastritis which was matched with normal mucosa. The expression level of PEBP1 in varioliform gastritis was significantly lower (P < 0.05) while that of TXNDC5 was significantly higher than that in matched normal gastric mucosa (P < 0.05). There are some changes of protein expression in varioliform gastritis. Downregulation of PEBP1 and upregulation of TXNDC5 are involved in the development of varioliform gastritis.

Comparative proteomic analysis of the esophageal squamous carcinoma cell line EC109 and its multi-drug resistant subline EC109/CDDP

To gain insights into the mechanisms of drug resistance in esophageal squamous cell carcinoma (ESCC), we employed proteomic techniques to study the global protein change of the multi-drug resistant ESCC cell line EC109/CDDP, which was established in our previous work, in comparison with its parental drug sensitive cell line EC109. By two-dimensional electrophoresis and mass spectrometry, we successfully identified 44 proteins with altered expression levels. These proteins are involved in endoplasmic reticulum stress response, metabolic process, DNA replication and repair, nucleotide binding, calcium binding, and cytoskeletal proteins. Among them, the differential expression levels of thioredoxin domain-containing protein 4 precursor and cystathionine gamma-lyase were further validated by Western blot and RT-PCR. Our present results lay foundation for future in-depth work on molecular mechanism of ESCC drug resistance, and aid in the identification and use of novel markers in clinical practice.

Differential Expression of Mimecan and Thioredoxin Domain–Containing Protein 5 in Colorectal Adenoma and Cancer: A Proteomic Study

Adenoma is the major precursor lesion of colorectal cancer, one of the most common cancers worldwide. The elucidation of the molecular mechanism underlying adenoma is essential for early detection, prevention, and intervention of colorectal cancer. Using a combination of two-dimensional gel electrophoresis and mass spectrometry, we identified 27 differentially expressed proteins in adenoma, compared with matched normal mucosa and cancer tissue. Seventeen proteins were upregulated and six downregulated in adenoma when compared with the same proteins in individual-matched normal mucosa. Four were downregulated, but none upregulated in adenoma when compared with the same proteins in matched cancer tissue. Two novel proteins, mimecan and thioredoxin domain-containing protein 5 (TXNDC5), were further validated by Western blot in 8 colorectal adenomas and 19 cancers that were matched with normal mucosa. All adenoma and cancer tissues did not express mimecan, but all normal mucosa did (P < 0.01). In contrast, TXNDC5 was significantly upregulated in colorectal adenoma and cancer tissues as compared with that in normal mucosa (P < 0.05). This study clearly demonstrated that absence of mimecan and upregulation of TXNDC5 are involved in the early development of colorectal cancer. Thus, the differentially expressed proteins might serve as potential biomarkers for colorectal cancer detection and intervention.

The neuronal differentiation process involves a series of antioxidant proteins

Involvement of individual antioxidant proteins (AOXP) and antioxidants in the differentiation process has been already reported. A systematic search strategy for detecting differentially regulated AOXP in neuronal differentiation, however, has not been published so far. The aim of this study was to provide an analytical tool identifying AOXP and to generate a differentiation-related AOXP expressional pattern. The undifferentiated N1E-115 neuroblastoma cell line was switched into a neuronal phenotype by DMSO treatment and used for proteomic experiments: We used two-dimensional gel electrophoresis followed by unambiguous mass spectrometrical (MALDI-TOF-TOF) identification of proteins to generate a map of AOXP. 16 AOXP were unambiguously determined in both cell lines; catalase, thioredoxin domain-containing protein 4 and hypothetical glutaredoxin/glutathione S-transferase C terminus-containing protein were detectable in the undifferentiated cells only. Five AOXP were observed in both, undifferentiated and differentiated cells and thioredoxin, thioredoxin-like protein p19, thioredoxin reductase 1, superoxide dismutases (Mn and Cu-Zn), glutathione synthetase, glutathione S-transferase P1 and Mu1 were detected in differentiated cells exclusively. Herein a differential expressional pattern is presented that reveals so far unpublished antioxidant principles involved in neuronal differentiation by a protein chemical approach, unambiguously identifying AOXP. This finding not only shows concomitant determination of AOXP but also serves as an analytical tool and forms the basis for design of future studies addressing AOXP and differentiation per se.