Translational Profiles of Medullary Myofibroblasts during Kidney Fibrosis
| Autor: | Andreas Hofmeister, Jing Liu, Andrew P. McMahon, Ivica Grgic, Benjamin D. Humphreys, Jeremy S. Duffield, A. Michaela Krautzberger, Derek P. DiRocco, Susanne V. Fleig, Matthew A. Lalli, Bruce J. Aronow |
|---|---|
| Rok vydání: | 2014 |
| Předmět: |
Ribosomal Proteins
Ribosomal Protein L10 Recombinant Fusion Proteins Transgene Green Fluorescent Proteins Mice Transgenic Biology Kidney Collagen Type I Mice Western blot Fibrosis Gene expression medicine Animals RNA Messenger Myofibroblasts Oligonucleotide Array Sequence Analysis Messenger RNA medicine.diagnostic_test Gene Expression Profiling Kidney metabolism General Medicine medicine.disease Molecular biology Up-Regulation Cell biology Collagen Type I alpha 1 Chain Mice Inbred C57BL Disease Models Animal Basic Research Real-time polymerase chain reaction Mice Inbred DBA Nephrology Myofibroblast Ureteral Obstruction |
| Zdroj: | Journal of the American Society of Nephrology. 25:1979-1990 |
| ISSN: | 1046-6673 |
| DOI: | 10.1681/asn.2013101143 |
| Popis: | Myofibroblasts secrete matrix during chronic injury, and their ablation ameliorates fibrosis. Development of new biomarkers and therapies for CKD will be aided by a detailed analysis of myofibroblast gene expression during the early stages of fibrosis. However, dissociating myofibroblasts from fibrotic kidney is challenging. We therefore adapted translational ribosome affinity purification (TRAP) to isolate and profile mRNA from myofibroblasts and their precursors during kidney fibrosis. We generated and characterized a transgenic mouse expressing an enhanced green fluorescent protein (eGFP)–tagged L10a ribosomal subunit protein under control of the collagen1α1 promoter. We developed a one-step procedure for isolation of polysomal RNA from collagen1α1-eGFPL10a mice subject to unilateral ureteral obstruction and analyzed and validated the resulting transcriptional profiles. Pathway analysis revealed strong gene signatures for cell proliferation, migration, and shape change. Numerous novel genes and candidate biomarkers were upregulated during fibrosis, specifically in myofibroblasts, and we validated these results by quantitative PCR, in situ, and Western blot analysis. This study provides a comprehensive analysis of early myofibroblast gene expression during kidney fibrosis and introduces a new technique for cell-specific polysomal mRNA isolation in kidney injury models that is suited for RNA-sequencing technologies. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|