LC-MS/MS identification of doublecortin as abundant beta-cell selective protein discharged by damaged beta cells in vitro
| Autor: | Zhidong Ling, F. K. Gorus, Peter Rotheneichner, Ludwig Aigner, Olivier Costa, C. Van Schravendijk, Sebastien Couillard-Despres, Geert Stangé, Benedicte Brackeva, K. Verhaeghen, Lei Jiang, Geert A. Martens, Daniel Pipeleers |
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| Přispěvatelé: | Diabetes Pathology & Therapy, Pathologic Biochemistry and Physiology, Pathology/molecular and cellular medicine |
| Předmět: |
Doublecortin Domain Proteins
Proteomics Doublecortin Protein Necrosis Biophysics Biochemistry Transcriptome In vivo Insulin-Secreting Cells Extracellular medicine Animals Humans Beta (finance) biology Glutamate Decarboxylase Neuropeptides Molecular biology In vitro Rats Doublecortin beta cells biology.protein medicine.symptom Beta cell Microtubule-Associated Proteins Biomarkers |
| Zdroj: | Vrije Universiteit Brussel |
| Popis: | There is a clinical need for plasma tests that can directly detect injury to pancreatic beta cells in type 1 diabetes. Such tests require biomarkers that are abundantly and selectively released into plasma by damaged beta cells. We combined LC-MS/MS proteomics and tissue-comparative transcriptomics of FACS-purified beta cells for bottom-up identification of candidate markers. Less than 10% of 467 proteins detected in beta cells showed endocrine-enriched expression. One surprising candidate was the neuronal migration marker doublecortin: in situ analysis revealed uniform doublecortin expression in the cytoplasm of all beta cells. Western blotting and real-time PCR confirmed its strong beta cell-selectivity outside the brain and its high molar abundance, indicating promising biomarker properties in comparison to GAD65, a more established marker of beta cell injury. DCX potential was validated in vitro: chemically-induced necrosis of rat and human beta cells led to a discharge of intracellular doublecortin into the extracellular space, proportionate to the amount of injured cells, and similar to GAD65. In vivo, recombinant DCX showed favorable pharmacokinetic properties, with a half-life in plasma of around 3h. Combined, our findings provide first proof-of-principle for doublecortin as biomarker for beta cell injury in vitro, advocating its further validation as biomarker in vivo. |
| Databáze: | OpenAIRE |
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