TMT-Based Quantitative Proteomics Reveals Cochlear Protein Profile Alterations in Mice with Noise-Induced Hearing Loss
Abstract
:1. Introduction
2. Materials and Methods
2.1. Animals and Noise-Induced Hearing Loss (NIHL) Model Construction
2.2. Auditory Brainstem Response (ABR) Measurement
2.3. Cochlea Surface Preparations and Hair Cells (HCs) Counting
2.4. Protein Extraction and Digestion
2.5. Tandem Mass Tag (TMT) Labeling and HPLC Fractionation
2.6. Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS) Analysis
2.7. Database Search and Bioinformatics Analysis
2.8. Oxidative Stress Markers Levels Assay, and Proinflammatory Cytokines Measurement
2.9. Western Blot Analysis
2.10. Statistical Analysis
3. Results
3.1. ABR Thresholds and HCs Immunofluorescence Staining and Counting
3.2. Identification of Differentially Expressed Proteins (DEPs)
3.3. Functional Enrichment Analysis of DEPs
3.4. Pathway Enrichment and Protein–Protein Interaction (PPI) Network Analysis
3.5. Validation of DEPs by Western Blot Analysis
3.6. Measurement of Superoxide Dismutase (SOD) Activity, Malondialdehyde (MDA) Level and Proinflammatory Cytokines Production
4. Discussion
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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KEGG Pathway | Protein | Count | p Value |
---|---|---|---|
PI3K/AKT signaling pathway | Col6a5, Prl, Gh1, Col1a1, Col2a1, Fgf1, Col1a2, Col9a2, Akt2, Itga1, Vwf | 11 | 0.033 |
ECM-receptor interaction | Col6a5, Col1a1, Col2a1, Dspp, Col1a2, Col9a2, Npnt, Itga1, Vwf | 9 | 9.34 × 10−6 |
Focal adhesion | Col6a5, Col1a1, Col2a1, Col1a2, Col9a2, Akt2, Mylpf, Itga1, Vwf | 9 | 0.005 |
Protein digestion and absorption | Col6a5, Col1a1, Col2a1, Col1a2, Col9a2, Col28a1, Col5a2, Col11a1, Col11a2, Slc38a2 | 10 | 2.11 × 10−6 |
Complement and coagulation cascades | Fga, A2m, Fgb, Fgg, Kng1, Serpina1a, Plg, Serpina1b, Serpina1e, Cfi, Vwf, C8g | 12 | 1.84 × 10−8 |
Platelet activation | Fga, Col1a1, Col1a2, Akt2, Fgb, Fgg, Vwf | 7 | 0.004 |
Neutrophil extracellular trap formation | Fga, Akt2, Hist2h2aa1, Fgb, Fgg, Vwf | 6 | 0.041 |
Nitrogen metabolism | Glul, Ca13 | 2 | 0.033 |
Longevity regulating pathway | Akt2, Atg5, Hspa1a, Cryab | 4 | 0.020 |
Selenocompound metabolism | Scly, Inmt | 2 | 0.033 |
Glycolysis/Gluconeogenesis | Hk2, Pgam2, Adh1, Eno3 | 4 | 0.022 |
Protein Accession | Protein Name | Protein Description | Molecular Weight (kDa) | p Value | Fold Change |
---|---|---|---|---|---|
Q3V3R4 | ITGA1 | Integrin alpha 1 | 130.81 | 0.013 | 1.22 |
O08677 | KNG1 | Kininogen 1 | 73.10 | 2.50 × 10−5 | 1.28 |
Q61129 | CFI | Complement factor I | 67.26 | 0.002 | 1.34 |
P61148 | FGF1 | Fibroblast growth factor 1 | 17.42 | 0.017 | 0.83 |
Q60823 | AKT2 | RAC-beta serine/threonine-protein kinase | 55.74 | 0.001 | 0.80 |
Q99J83 | ATG5 | Autophagy protein 5 | 32.40 | 0.019 | 0.78 |
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Miao, L.; Zhang, J.; Yin, L.; Pu, Y. TMT-Based Quantitative Proteomics Reveals Cochlear Protein Profile Alterations in Mice with Noise-Induced Hearing Loss. Int. J. Environ. Res. Public Health 2022, 19, 382. https://doi.org/10.3390/ijerph19010382
Miao L, Zhang J, Yin L, Pu Y. TMT-Based Quantitative Proteomics Reveals Cochlear Protein Profile Alterations in Mice with Noise-Induced Hearing Loss. International Journal of Environmental Research and Public Health. 2022; 19(1):382. https://doi.org/10.3390/ijerph19010382
Chicago/Turabian StyleMiao, Long, Juan Zhang, Lihong Yin, and Yuepu Pu. 2022. "TMT-Based Quantitative Proteomics Reveals Cochlear Protein Profile Alterations in Mice with Noise-Induced Hearing Loss" International Journal of Environmental Research and Public Health 19, no. 1: 382. https://doi.org/10.3390/ijerph19010382
APA StyleMiao, L., Zhang, J., Yin, L., & Pu, Y. (2022). TMT-Based Quantitative Proteomics Reveals Cochlear Protein Profile Alterations in Mice with Noise-Induced Hearing Loss. International Journal of Environmental Research and Public Health, 19(1), 382. https://doi.org/10.3390/ijerph19010382