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Enrichment of Extracellular Matrix Proteins from Tissues and Digestion into Peptides for Mass Spectrometry Analysis
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Quantitative extracellular matrix proteomics to study mammary and liver tissue microenvironments
Erica T Goddard1, Ryan C Hill2, Alexander Barrett2
1Department of Cell, Developmental and Cancer Biology, Oregon Health & Science University, Portland, OR, USA.
The International Journal of Biochemistry & Cell Biology
|October 30, 2016
Summary
The extracellular matrix (ECM) in the mammary gland and liver has distinct compositions. ECM changes during the postpartum period may increase breast cancer metastasis risk.
Area of Science:
- Biochemistry
- Cell Biology
- Proteomics
Background:
- The extracellular matrix (ECM) is crucial for epithelial cell function, tissue homeostasis, and disease, including cancer.
- Understanding tissue-specific ECM composition and its role in cancer progression is limited.
- Existing proteomic methods struggle to quantify absolute ECM protein abundance.
Purpose of the Study:
- To develop and apply a mass spectrometry-based method for absolute quantification of ECM proteins in breast cancer-relevant tissues.
- To compare ECM composition and abundance between the normal mammary gland and liver.
- To investigate ECM changes during the reproductive cycle in the mammary gland and their potential link to cancer risk.
Main Methods:
- Utilized mass spectrometry optimized for absolute quantitative ECM analysis.
- Employed an extensive protein extraction and digestion method.
- Incorporated stable isotope labeled Quantitative conCATamer (QconCAT) peptides as internal standards for absolute protein quantification.
- Analyzed 98 ECM, ECM-associated, and cellular proteins in a single run.
Main Results:
- Quantified 98 ECM and related proteins in rodent mammary gland and liver tissues.
- Identified distinct ECM abundance and relative composition between mammary gland and liver.
- Observed significant ECM compositional changes in the mammary gland during weaning-induced involution, a pro-tumorigenic window.
- Demonstrated differential ECM profiles across the reproductive cycle in the mammary gland.
Conclusions:
- The study provides a quantitative proteomic approach to analyze tissue-specific ECM.
- Distinct ECM profiles in mammary gland and liver suggest tissue-specific roles in metastasis.
- Postpartum-associated ECM changes, particularly during involution, may represent a critical window for breast cancer metastasis.
- Identified potential ECM candidates for further investigation in breast cancer progression and metastasis.
- The developed method can enable the creation of more accurate in vitro models for studying tumor-ECM dynamics.
Keywords:
Breast cancerExtracellular matrixLiverLiver metastasisMammary glandMass spectrometry proteomicsMore Related Videos
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