A serologically assessed neo-epitope biomarker of cellular fibronectin degradation is related to pulmonary fibrosis
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Background: Idiopathic pulmonary fibrosis (IPF) is characterized by excessive extracellular matrix (ECM) remodeling, herein ECM degradation. Fibronectin (FN) is an important component of the ECM that is produced by multiple cell types, including fibroblasts. Extra domain B (EDB) is specific for a cellular FN isoform which is found in the ECM. We sought to develop a non-invasive test to investigate whether matrix metalloproteinase 8 (MMP-8) degradation of EDB in cellular FN results in a specific protein fragment that can be assessed serologically and if levels relate to pulmonary fibrosis. Method: Cellular FN was cleaved in vitro by MMP-8 and a protein fragment was identified by mass spectrometry. A monoclonal antibody (mAb) was generated, targeting a neo-epitope originating from EDB in cellular FN. Utilizing this mAb, a neo-epitope specific enzyme-linked immunosorbent assay (FN-EDB) was developed and technically validated. Serum FN-EDB was assessed in an IPF cohort (n = 98), registered at clinicaltrials.gov (NCT02818712), and in healthy controls (n = 35). Results: The FN-EDB assay had high specificity for the MMP-8 degraded neo-epitope and was technically robust. FN-EDB serum levels were not influenced by age, sex, ethnicity, or BMI. Moreover, FN-EDB serum levels were significantly higher in IPF patients (median 31.38 [IQR 25.79–46.84] ng/mL) as compared to healthy controls (median 28.05 [IQR 21.58–33.88] ng/mL, p = 0.023). Conclusion: We developed the neo-epitope specific FN-EDB assay, a competitive ELISA, as a tool for serological assessment of MMP-8 mediated degradation of EDB in cellular FN. This study indicates that degradation of EDB in cellular FN is elevated in IPF and warrants further investigation.
| Originalsprog | Engelsk |
|---|---|
| Artikelnummer | 110599 |
| Tidsskrift | Clinical Biochemistry |
| Vol/bind | 118 |
| Antal sider | 8 |
| ISSN | 0009-9120 |
| DOI | |
| Status | Udgivet - 2023 |
Bibliografisk note
Funding Information:
The Q-Exactive mass spectrometer was granted by Independent Research Fund Denmark | Natural Sciences (ID: DFF – 1335-00071 to BS). Moreover, this study has been funded by the Danish Research Foundation, TrygFonden (grant 118860), Aarhus University (unrestricted), Boehringer Ingelheim Denmark (unrestricted), the Danish Lung Association’s Fund, the Health Research Fund of the Central Denmark Region, and the Ellen and Knud Dalhoff Larsen’s Fund. The funders of the study had no role in study design, data collection, data analyses, data interpretation, writing of the report or decision to submit the paper for publication.
Funding Information:
The authors wish to acknowledge everyone who has been involved in the work for this study. This includes participants, medical-, nursing-, and technical-staff involved in acquiring and supplying samples and measurements. Additionally, we wish to acknowledge Lene Holberg Blicher for their help with mass spectrometry measurements and analysis. Lastly, we wish to acknowledge all the organizations who funded this study. The data obtained in the current study are included in this published article, available upon request. The Q-Exactive mass spectrometer was granted by Independent Research Fund Denmark | Natural Sciences (ID: DFF – 1335-00071 to BS). Moreover, this study has been funded by the Danish Research Foundation, TrygFonden (grant 118860), Aarhus University (unrestricted), Boehringer Ingelheim Denmark (unrestricted), the Danish Lung Association's Fund, the Health Research Fund of the Central Denmark Region, and the Ellen and Knud Dalhoff Larsen's Fund. The funders of the study had no role in study design, data collection, data analyses, data interpretation, writing of the report or decision to submit the paper for publication. Study concept and design: AHH, HWB, DGKR, FG, JMBS, DJL. Acquisition of data: AHH, HWB, TSP, DGKR, MVL, BS, LLL, TMJ, EB. Analysis and interpretation of data: AHH, HWB. Experimental support: JMBS, DJL. Drafting of the manuscript: AHH, HWB. Critical revision of the manuscript for important intellectual content: AHH, HWB, TSP, OH, DGKR, FG, MVL, BS, LLL, TMJ, MAK, DJL, EB, JMBS.
Publisher Copyright:
© 2023 The Authors
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