Abstract
BackgroundPituitary adenomas, the third most common intracranial tumor, comprise nearly 16.7% of intracranial neoplasm and 25%-44% of pituitary adenomas are prolactinomas. Prolactinoma represents a complex heterogeneous mixture of cells including prolactin (PRL), endothelial cells, fibroblasts, and other stromal cells, making it difficult to dissect the molecular and cellular mechanisms of prolactin cells in pituitary tumorigenesis through high-throughout-omics analysis. Our newly developed immuno-laser capture microdissection (LCM) method would permit rapid and reliable procurement of prolactin cells from this heterogeneous tissue. Thus, prolactin cell specific molecular events involved in pituitary tumorigenesis and cell signaling can be approached by proteomic analysis.ResultsProteins from immuno-LCM captured prolactin cells were digested; resulting peptides were separated by two dimensional-nanoscale liquid chromatography (2D-nanoLC/MS) and characterized by tandem mass spectrometry. All MS/MS spectrums were analyzed by SEQUEST against the human International Protein Index database and a specific prolactinoma proteome consisting of 2243 proteins was identified. This collection of identified proteins by far represents the largest and the most comprehensive database of proteome for prolactinoma. Category analysis of the proteome revealed a widely unbiased access to various proteins with diverse functional characteristics.ConclusionsThis manuscript described a more comprehensive proteomic profile of prolactinomas compared to other previous published reports. Thanks to the application of immuno-LCM combined with online two-dimensional nano-scale liquid chromatography here permitted identification of more proteins and, to our best knowledge, generated the largest prolactinoma proteome. This enlarged proteome would contribute significantly to further understanding of prolactinoma tumorigenesis which is crucial to the management of prolactinomas.
Highlights
Prolactinomas are the most common pituitary tumors, representing 25%-44% of all pituitary adenoma cases [1]
laser capture microdissection (LCM) allows the isolation of even single cell and immunohistochemistry (IHC) staining could help to distinguish specific cell populations, employment of immuno-LCM could pick up certain cell populations with specific immuno-phenotype from complex tissues according to their antigen expression to ameliorate the problem of tissue heterogeneity in proteomic research [5,6]
In a typical shotgun proteomics experiment, a complex protein sample is digested into peptides followed by separation by 2D-nanoLC/mass spectrometry (MS), resulting peptides were loaded into a mass spectrometer for fragmentation and sequencing
Summary
Prolactinomas are the most common pituitary tumors, representing 25%-44% of all pituitary adenoma cases [1]. While 2-DE serves as a powerful separation tool, shotgun proteomics combining LC-MS/MS have emerged as a technique of choice for large-scale protein studies due to its superior throughput and sensitivity [8]. In a typical shotgun proteomics experiment, a complex protein sample is digested into peptides followed by separation by 2D-nanoLC/MS, resulting peptides were loaded into a mass spectrometer for fragmentation and sequencing. High performance duty cycle instruments can fragment hundreds of peptides in a single LC-MS/MS experiment. Prolactinoma represents a complex heterogeneous mixture of cells including prolactin (PRL), endothelial cells, fibroblasts, and other stromal cells, making it difficult to dissect the molecular and cellular mechanisms of prolactin cells in pituitary tumorigenesis through high-throughoutomics analysis. Prolactin cell specific molecular events involved in pituitary tumorigenesis and cell signaling can be approached by proteomic analysis
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