Proteomic analysis of the proliferative and secretory phases of the human endometrium: Protein identification and differential protein expression

DeSouza, Leroi V.; Diehl, Georg; Yang, Eric C.; Guo, Jingzhong; Rodrigues, Mary Joe; Romaschin, Alexander D.; Colgan, Terence J.; Siu, Kin Wai Michael

doi:10.1002/pmic.200400920

Cited by 97 publications

(94 citation statements)

References 26 publications

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“…Two hundred micrograms of each of the four GBM lysates were digested and labeled with the following iTRAQ labels: wtEGFR, 114 and 116 labels; EGFRvIII, 115 and 117 labels. Samples were processed by offline two-dimensional LC-MS/MS analysis as described previously (19,20). After separation by strong cation exchange (SCX), each fraction was dried and resuspended in 10 μL of 0.1% formic acid.…”

Section: Methodsmentioning

confidence: 99%

“…The development of novel mass spectrometry (MS)-based technologies, such as iTRAQ-labeling, yields great promise in our ability to interrogate differences in the proteome between pathologic and normal tissues (19)(20)(21). In the present investigation, we used preclinical in vitro and in vivo human GBM models that are well characterized for their expression profiles of wtEGFR and EGFRvIII.…”

Section: Introductionmentioning

confidence: 99%

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Loss of Collapsin Response Mediator Protein1, as Detected by iTRAQ Analysis, Promotes Invasion of Human Gliomas Expressing Mutant EGFRvIII

et al. 2009

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Glioblastoma multiforme (GBM) is the most common and lethal primary human brain tumor. GBMs are characterized by a variety of genetic alterations, among which oncogenic mutations of epidermal growth factor receptor (EGFRvIII) is most common. GBMs harboring EGFRvIII have increased proliferation and invasive characteristics versus those expressing wildtype (wt) EGFR. To identify the molecular basis of this increased tumorgenic phenotype, we used iTRAQ-labeling differential proteomic analysis. Among several differentially expressed proteins, we selected CRMP1, a protein implicated in cellular invasion that was markedly decreased in GBMs expressing EGFRvIII, for further study. The differential expression of CRMP1 was confirmed in a panel of human GBM cell lines and operative specimens that express wtEGFR or mutant EGFRvIII by quantitative real-time PCR, Western blot, and immunohistochemical analysis. In human GBM samples, decreased expression of CRMP1 correlated with EGFRvIII positivity. Knockdown of CRMP1 by siRNA resulted in increased invasion of wtEGFR expressing human GBM cells (U87 and U373) to those found in isogenic GBM cells. Exogenous expression of EGFRvIII in these wtEGFR-expressing GBM cells promoted their ability to invade and was accompanied by decreased expression of CRMP1. Rescuing CRMP1 expression decreased invasion of the EGFRvIII-expressing GBM cells by tilting the balance between Rac and Rho. Collectively, these results show that the loss of CRMP1 contribute to the increased invasive phenotype of human GBMs expressing mutant EGFRvIII. [Cancer Res 2009;69(22):8545-54]

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Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Loss of Collapsin Response Mediator Protein1, as Detected by iTRAQ Analysis, Promotes Invasion of Human Gliomas Expressing Mutant EGFRvIII

et al. 2009

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“…Our recent studies using both ICAT and iTRAQ reagents as means to facilitate the identification and relative quantification of proteins from endometrial tissue homogenates have resulted in some interesting potential cancer markers (PCMs) (2,3). Those studies, however, were performed on small sample sets.…”

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confidence: 99%

Endometrial Carcinoma Biomarker Discovery and Verification Using Differentially Tagged Clinical Samples with Multidimensional Liquid Chromatography and Tandem Mass Spectrometry

DeSouza

Grigull

Ghanny

et al. 2007

Molecular & Cellular Proteomics

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. (2005) Search for cancer markers from endometrial tissues using differentially labeled tags iTRAQ and cleavable ICAT with multidimensional liquid chromatography and tandem mass spectrometry. J. Proteome Res. 4, 377-386) to discriminate malignant and benign endometrial tissue samples was verified in a 40-sample iTRAQ (isobaric tags for relative and absolute quantitation) labeling study involving normal proliferative and secretory samples and Types I and II endometrial cancer samples. None of these proteins had the sensitivity and specificity to be used individually to discriminate between normal and cancer samples. However, a panel of pyruvate kinase, chaperonin 10, and ␣ 1 -antitrypsin achieved the best results with a sensitivity, specificity, predictive value, and positive predictive value of 0.95 each in a logistic regression analysis. In addition, three new potential markers were discovered, whereas two other proteins showed promising trends but were not detected in sufficient numbers of samples to permit statistical validation. Differential expressions of some of these candidate biomarkers were independently verified using immunohistochemistry. Molecular & Cellular Proteomics 6:1170 -1182, 2007.Differential tagging with isotopic reagents, such as ICAT (1) or the more recent variation that uses isobaric tagging reagents, iTRAQ 1 (Applied Biosystems, Foster City, CA), followed by multidimensional LC and MS/MS analysis is quickly being recognized as one of the more powerful methodologies in the search for biomarkers for various disease states. Our recent studies using both ICAT and iTRAQ reagents as means to facilitate the identification and relative quantification of proteins from endometrial tissue homogenates have resulted in some interesting potential cancer markers (PCMs) (2, 3). Those studies, however, were performed on small sample sets. This study describes the results of a more detailed investigation using a larger cohort of 40 samples and the iTRAQ technology and was aimed at validating the earlier results as well as expanding the panel of biomarkers.Endometrial carcinoma (EmCa), a cancer of the lining of the uterus, is the fourth most common cancer in Canadian women.2 Current methods of diagnosis rely on invasive techniques (biopsy and curettage), and no screening is available. A panel of biomarkers that helps in early diagnosis would, therefore, be useful especially for high risk groups, e.g. women who are on tamoxifen treatment or have hereditary nonpolyposis colorectal cancer syndrome. Although the eventual diagnostic testing for such biomarkers would be most facile from bodily fluids, such as blood or urine, the iTRAQ experiments performed thus far have been on resected From the Departments of ‡Chemistry, §Biology, and ʈMathematics

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“…This is due to posttranscriptional regulatory mechanisms (mRNA export, surveillance, silencing and turnover) and posttranslational modifications, all of which can determine protein activity, localisation, turnover and interactions with other proteins (Mann & Jensen 2003). Several studies have previously sought to identify proteins differentially expressed in the human endometrium between the proliferative and secretory phases of normal menstrual cycles (DeSouza et al 2005, Chen et al 2009, Domínguez et al 2009, Parmar et al 2009, Rai et al 2010. There are also a number of studies describing endometrial phasespecific transcriptomic profiles (Kao et al 2002, Ace & Okulicz 2004, Evans et al 2012, Garrido-Gómez et al 2013.…”

Section: Introductionmentioning

confidence: 99%

Proteomic analysis of the sheep caruncular and intercaruncular endometrium reveals changes in functional proteins crucial for the establishment of pregnancy

Al-Gubory¹,

Arianmanesh²,

Garrel³

et al. 2014

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The expression and regulation of endometrial proteins are crucial for conceptus implantation and development. However, little is known about site-specific proteome profiles of the mammalian endometrium during the peri-implantation period. We utilised a two-dimensional gel electrophoresis/mass spectrometry-based proteomics approach to compare and identify differentially expressed proteins in sheep endometrium. Caruncular and intercaruncular endometrium were collected on days 12 (C12) and 16 (C16) of the oestrous cycle and at three stages of pregnancy corresponding to conceptus pre-attachment (P12), implantation (P16) and post-implantation (P20). Abundance and localisation changes in differentially expressed proteins were determined by western blot and immunohistochemistry. In caruncular endometrium, 45 protein spots (5% of total spots) altered between day 12 of pregnancy (P12) and P16 while 85 protein spots (10% of total spots) were differentially expressed between P16 and C16. In intercaruncular endometrium, 31 protein spots (2% of total spots) were different between P12 and P16 while 44 protein spots (4% of total spots) showed differential expression between C12 and C16. The pattern of protein changes between caruncle and intercaruncle sites was markedly different. Among the protein spots with implantation-related changes in volume, 11 proteins in the caruncular endometrium and six proteins in the intercaruncular endometrium, with different functions such as protein synthesis and degradation, antioxidant defence, cell structural integrity, adhesion and signal transduction, were identified. Our findings highlight the different but important roles of the caruncular and intercaruncular proteins during early pregnancy.

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Proteomic analysis of the proliferative and secretory phases of the human endometrium: Protein identification and differential protein expression

Cited by 97 publications

References 26 publications

Loss of Collapsin Response Mediator Protein1, as Detected by iTRAQ Analysis, Promotes Invasion of Human Gliomas Expressing Mutant EGFRvIII

Loss of Collapsin Response Mediator Protein1, as Detected by iTRAQ Analysis, Promotes Invasion of Human Gliomas Expressing Mutant EGFRvIII

Endometrial Carcinoma Biomarker Discovery and Verification Using Differentially Tagged Clinical Samples with Multidimensional Liquid Chromatography and Tandem Mass Spectrometry

Proteomic analysis of the sheep caruncular and intercaruncular endometrium reveals changes in functional proteins crucial for the establishment of pregnancy

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