Comparative Proteomic Profiling of Secreted Extracellular Vesicles from Breast Fibroadenoma and Malignant Lesions: A Pilot Study
Abstract
:1. Introduction
2. Materials and Methods
2.1. Cell Lines and Primary Cultures
2.2. Purification of Extracellular Vesicles
2.3. Dynamic Light Scattering (DLS) Measurements
2.4. Scanning Electron Microscopy (SEM)
2.5. Liquid Chromatography Tandem Mass Spectrometry (LC-MS/MS)
2.6. Database Searches and Bioinformatics Analyses
2.7. Western Blot Analysis
3. Results
3.1. Study Design and Sample Description
3.2. Characterization of EVs from Primary and IBCC Lines
Patient | Sample #37 | Sample #46 | Sample #72 | Sample #44 | Sample #170 | Sample #148 |
---|---|---|---|---|---|---|
Median age at diagnosis (years) | 49.5 | 43 | 53.5 | |||
Age at diagnosis (years) | 66 | 33 | 44 | 42 | 56 | 51 |
Histological type | Ductal infiltrating carcinoma (NOS) | Ductal infiltrating carcinoma high grade | Fibroadenoma | Fibroadenoma with adenosi | Ductal infiltrating carcinoma (NOS) CK19 (+++) | Lobular infiltrating carcinoma, poorly differentiated with 10% of lobular neoplasia in situ with high grade, E-cadherin negative |
Tumor stage | pT1c | pT2 | _ | _ | pT2 | pT2 |
Grade | pG3 | pG3 | _ | _ | pG3 | pG3 |
Lymph node | pN1a | pN1a | _ | _ | 0 | 0 |
ER/PR/HER2 status (positivity) | ER − (0)/PR + (<5)/HER2 + (2+) | ER +(10)/PR + a/HER2 + (3+) | _ | _ | ER − (0)/PR − (0)/HER2 − (0) | ER − (0)/PR − (<5)/HER2 − (0) |
ki 67 status (positivity) | High (30) | High (50) | _ | _ | High (80) | High (45) |
Subtype | HER2+ BC | HER2+ BC | FAD | FAD | TNBC | TNBC |
Corresponding cell model | BT-549 | MCF10-A | MDA-MB-231 |
3.3. Qualitative and Semi-Quantitative MS-Based Proteomic Profiling
3.4. Pathway Analysis
3.5. Protein Abundance Patterns and In Vitro Investigation
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
Abbreviations
BC | breast cancer |
DDM | n-dodecyl β-D-maltoside |
DLS | dynamic light scattering |
ESI | electrospray ionization |
ER | estrogen receptor |
EVs | extracellular vesicles |
FAD | fibroadenoma |
FDR | false discovery rate |
HER2 | human epidermal growth factor receptor 2 |
IBCC | immortalized breast cell culture |
PR | progesterone receptor |
SED | secondary electron detector |
SEM | scanning electron microscopy |
TNBC | triple–negative breast cancer |
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Sample | Mean Diameter (nm) ± S.D. | PDI | (D ± S.D.) * 10−12 m2 s−1 |
---|---|---|---|
#37 | 346 ± 70 | 0.356 | 1.15 ± 0.23 |
#44 | 270 ± 74 8 ± 2 | 0.395 | 1.47 ± 0.40 49.6 ± 12.4 |
#148 | 298 ± 64 | 0.365 | 1.33 ± 0.29 |
MDA-MB-231 | 295 ± 58 | 0.310 | 1.39 ± 0.27 |
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Pane, K.; Quintavalle, C.; Nuzzo, S.; Ingenito, F.; Roscigno, G.; Affinito, A.; Scognamiglio, I.; Pattanayak, B.; Gallo, E.; Accardo, A.; et al. Comparative Proteomic Profiling of Secreted Extracellular Vesicles from Breast Fibroadenoma and Malignant Lesions: A Pilot Study. Int. J. Mol. Sci. 2022, 23, 3989. https://doi.org/10.3390/ijms23073989
Pane K, Quintavalle C, Nuzzo S, Ingenito F, Roscigno G, Affinito A, Scognamiglio I, Pattanayak B, Gallo E, Accardo A, et al. Comparative Proteomic Profiling of Secreted Extracellular Vesicles from Breast Fibroadenoma and Malignant Lesions: A Pilot Study. International Journal of Molecular Sciences. 2022; 23(7):3989. https://doi.org/10.3390/ijms23073989
Chicago/Turabian StylePane, Katia, Cristina Quintavalle, Silvia Nuzzo, Francesco Ingenito, Giuseppina Roscigno, Alessandra Affinito, Iolanda Scognamiglio, Birlipta Pattanayak, Enrico Gallo, Antonella Accardo, and et al. 2022. "Comparative Proteomic Profiling of Secreted Extracellular Vesicles from Breast Fibroadenoma and Malignant Lesions: A Pilot Study" International Journal of Molecular Sciences 23, no. 7: 3989. https://doi.org/10.3390/ijms23073989