한빛사논문
- 조회506
Nhuan T. Do1,2†, Sun Young Lee1†, Yoon Seo Lee1†, ChaeHo Shin3,4, Daeho Kim5, Tae Geol Lee1,4, Jin Gyeong Son1* and Se‑Hwa Kim1,2*
1Safety Measurement Institute, Korea Research Institute of Standards and Science, 267 Gajeong‑Ro, Yuseong‑Gu, Daejeon 34113, Republic of Korea.
2BioMedical Measurement, University of Science and Technology, 217 Gajeong‑Ro, Yuseong‑Gu, Daejeon 34113, Republic of Korea.
3Interdisciplinary Materials Measurement Institute, Korea Research Institute of Standards and Science, 267 Gajeong‑Ro, Yuseong‑Gu, Daejeon 34113, Republic of Korea.
4Nanoconvergence Measurement, University of Science and Technology, 217 Gajeong‑Ro, Yuseong‑Gu, Daejeon 34113, Republic of Korea.
5Bruker Nano Surface & Metrology, Bruker Korea, Seongnam 13493, Republic of Korea.
†Nhuan T. Do, Sun Young Lee and Yoon Seo Lee contributed equally to this work.
Corresponding authors : Correspondence to Jin Gyeong Son or Se-Hwa Kim.
Abstract
Background: Fibrosis plays an important role in both normal physiological and pathological phenomena as fibroblasts differentiate to myofibroblasts. The activation of fibroblasts is determined through interactions with the surrounding extracellular matrix (ECM). However, how this fibroblast-to-myofibroblast transition (FMT) is regulated and affected by elastin concentration in a three-dimensional (3D) microenvironment has not been investigated.
Methods: We developed an insoluble elastin-gradient 3D hydrogel system for long-lasting cell culture and studied the molecular mechanisms of the FMT in embedded cells by nanoflow LC-MS/MS analysis along with validation through real-time PCR and immunofluorescence staining.
Results: By optimizing pH and temperature, four 3D hydrogels containing fibroblasts were successfully fabricated having elastin concentrations of 0, 20, 50, and 80% in collagen. At the low elastin level (20%), fibroblast proliferation was significantly increased compared to others, and in particular, the FMT was clearly observed in this condition. Moreover, through mass spectrometry of the hydrogel environment, it was confirmed that differentiation proceeded in two stages. In the early stage, calcium-dependent proteins including calmodulin and S100A4 were highly associated. On the other hand, in the late stage after several passages of cells, distinct markers of myofibroblasts were presented such as morphological changes, increased production of ECM, and increased α-SMA expression. We also demonstrated that the low level of elastin concentration induced some cancer-associated fibroblast (CAF) markers, including PDGFR-β, and fibrosis-related disease markers, including THY-1.
Conclusion: Using our developed 3D elastin-gradient hydrogel system, we evaluated the effect of different elastin concentrations on the FMT. The FMT was induced even at a low concentration of elastin with increasing CAF level via calcium signaling. With this system, we were able to analyze varying protein expressions in the overall FMT process over several cellular passages. Our results suggest that the elastin-gradient system employing nonlinear optics imaging provides a good platform to study activated fibroblasts interacting with the microenvironment, where the ECM plays a pivotal role.
논문정보
- Research article
- 2023년 10월 (BRIC 등록일 2023-10-19)
- 국내 연구진
- 바이오·의료융합 > 다학제 융합/복합
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