Analysis of mouse brain microvascular endothelium using immuno‐laser capture microdissection coupled to a hybrid linear ion trap with Fourier transform‐mass spectrometry proteomics platform

Lu, Qiaozhen; Murugesan, Nivetha; Macdonald, Jennifer A.; Wu, Shiaw‐Lin; Pachter, Joel S.; Hancock, William S.

doi:10.1002/elps.200700936

Cited by 38 publications

(26 citation statements)

References 33 publications

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“…Laser capture microdissection is often used in conjunction with MS-based protein identification technology to assist with the discovery of tumor-associated molecules in tissue specimens containing various types of cells (27,31,(61)(62)(63)(64). However, relatively few studies have used these techniques to identify OSCC/HNSCC-associated proteins in tissue specimens from patients with OSCC/HNSCC (15,17,29).…”

Section: Discussionmentioning

confidence: 99%

Enhanced Interferon Signaling Pathway in Oral Cancer Revealed by Quantitative Proteome Analysis of Microdissected Specimens Using 16O/18O Labeling and Integrated Two-dimensional LC-ESI-MALDI Tandem MS

Chi

Lee

Chang

et al. 2009

Molecular & Cellular Proteomics

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Section: Discussionmentioning

confidence: 99%

Enhanced Interferon Signaling Pathway in Oral Cancer Revealed by Quantitative Proteome Analysis of Microdissected Specimens Using 16O/18O Labeling and Integrated Two-dimensional LC-ESI-MALDI Tandem MS

Chi

Lee

Chang

et al. 2009

Molecular & Cellular Proteomics

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“…17,24 However, to our knowledge, temporal proteomic profiles of human brain endothelial responses have not been reported. Differences between human and rodent cells are not fully understood.…”

Section: Discussionmentioning

confidence: 99%

Proteomic Temporal Profile of Human Brain Endothelium After Oxidative Stress

Ning

Sarracino

Kho

et al. 2011

Stroke

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Background and Purpose Because brain endothelial cells exist at the neurovascular interface, they may serve as cellular reporters of brain dysfunction by releasing biomarkers into the circulation. Methods We used proteomic techniques to screen conditioned media from human brain endothelial cultures subjected to oxidative stress induced by nitric oxide over 24 hours. Plasma samples from human stroke patients were analyzed by enzyme-linked immunosorbent assay. Results In healthy endothelial cells, interaction mapping demonstrated cross-talk involving secreted factors, membrane receptors, and matrix components. In oxidatively challenged endothelial cells, networks of interacting proteins failed to emerge. Instead, inflammatory markers increased, secreted factors oscillated over time, and endothelial injury repair was manifested as changes in factors related to matrix integrity. Elevated inflammatory markers included heat shock protein, chemokine ligand-1, serum amyloid-A1, annexin-A5, and thrombospondin-1. Neurotrophic factors (prosaposin, nucleobindin-1, and tachykinin precursors) peaked at 12 hours, then rapidly decreased by 24 hours. Basement membrane components (fibronectin, desomoglein, profiling-1) were decreased. Cytoskeletal markers (actin, vimentin, nidogen, and filamin B) increased over time. From this initial analysis, the high-ranking candidate thrombospondin-1 was further explored in human plasma. Acute ischemic stroke patients had significantly higher thrombospondin-1 levels within 8 hours of symptom onset compared to controls with similar clinical risk factors (659±81 vs 1132±98 ng/mL; P<0.05; n=20). Conclusions Screening of simplified cell culture systems may aid the discovery of novel biomarkers in clinical neurovascular injury. Further collaborative efforts are warranted to discover and validate more candidates of interest.

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“…However, the quantity of recovered protein varies depending on the method used for immunostaining, colorimetric versus fluorescent. Other laboratories have had similar observations and switched from a horseradish peroxidase-based approach to an alkaline phosphatase system (Lu et al 2008). At present, use of fluorescent staining and auto-recognition software is the preferred staining approach (Fig.…”

Section: Protein Analysismentioning

confidence: 91%

Effect of Immunohistochemistry on Molecular Analysis of Tissue Samples

Tangrea

Mukherjee

Gao

et al. 2011

J Histochem Cytochem.

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The field of molecular pathology has advanced over the past 15 years with the integration of laser-based microdissection systems and a variety of associated biological assays and analysis methods (Bonner et al. 1997; EmmertBuck et al. 1996;Burgess and Hazelton 2000;Iyer and Cox 2010;Craven and Banks 2001;Mustafa et al. 2008;Lawrie and Curran 2005;Charboneau et al. 2002;Gillespie et al. 2004;Bichsel et al. 2001;Callagy et al. 2005;Kurihara et al. 2002) SummaryLaser-based tissue microdissection is an important tool for the molecular evaluation of histological sections. The technology has continued to advance since its initial commercialization in the 1990s, with improvements in many aspects of the process. More recent developments are tailored toward an automated, operator-independent mode that relies on antibodies as targeting probes, such as immuno-laser capture microdissection or expression microdissection (xMD). Central to the utility of expression-based dissection techniques is the effect of the staining process on the biomolecules in histological sections. To investigate this issue, the authors analyzed DNA, RNA, and protein in immunostained, microdissected samples. DNA was the most robust molecule, exhibiting no significant change in quality after immunostaining but a variable 50% to 75% decrease in the total yield. In contrast, RNA in frozen and ethanol-fixed, paraffin-embedded samples was susceptible to hydrolysis and digestion by endogenous RNases during the initial steps of staining. Proteins from immunostained tissues were successfully analyzed by one-dimensional electrophoresis and mass spectrometry but were less amenable to solution phase assays. Overall, the results suggest investigators can use immunoguided microdissection methods for important analytic techniques; however, continued improvements in staining protocols and molecular extraction methods are key to further advancing the capability of these methods. (J Histochem Cytochem 59:591-600, 2011)

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Analysis of mouse brain microvascular endothelium using immuno‐laser capture microdissection coupled to a hybrid linear ion trap with Fourier transform‐mass spectrometry proteomics platform

Cited by 38 publications

References 33 publications

Enhanced Interferon Signaling Pathway in Oral Cancer Revealed by Quantitative Proteome Analysis of Microdissected Specimens Using 16O/18O Labeling and Integrated Two-dimensional LC-ESI-MALDI Tandem MS

Enhanced Interferon Signaling Pathway in Oral Cancer Revealed by Quantitative Proteome Analysis of Microdissected Specimens Using 16O/18O Labeling and Integrated Two-dimensional LC-ESI-MALDI Tandem MS

Proteomic Temporal Profile of Human Brain Endothelium After Oxidative Stress

Effect of Immunohistochemistry on Molecular Analysis of Tissue Samples

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