Proteomic Analysis of Nucleus Pulposus Cell-derived Extracellular Matrix Niche and Its Effect on Phenotypic Alteration of Dermal Fibroblasts

Minting Yuan,Barbara P Chan,Henry Lam,Pei-Jing Pai,Xiaofen Liu

doi:10.1038/s41598-018-19931-9

Minting Yuan, Barbara P Chan + Show 3 more

Open Access

https://doi.org/10.1038/s41598-018-19931-9

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Abstract

Reconstituting biomimetic matrix niche in vitro and culturing cells at the cell niche interface is necessary to understand the effect and function of the specific matrix niche. Here we attempted to reconstitute a biomimetic extracellular matrix (ECM) niche by culturing nucleus pulposus cells (NPCs) in a collagen microsphere system previously established and allowing them to remodel the template matrix. The reconstituted NPC-derived complex ECM was obtained after decellularization and the composition of such niche was evaluated by proteomic analysis. Results showed that a complex acellular matrix niche consisting of ECM proteins and cytoskeletal proteins by comparing with the template collagen matrix starting material. In order to study the significance of the NPC-derived matrix niche, dermal fibroblasts were repopulated in such niche and the phenotypes of these cells were changed, gene expression of collagen type II and CA12 increased significantly. A biomimetic NPC-derived cell niche consisting of complex ECM can be reconstituted in vitro, and repopulating such matrix niche with fibroblasts resulted in changes in phenotypic markers. This work reports a 3D in vitro model to study cell niche factors, contributing to future understanding of cellular interactions at the cell-niche interface and rationalized scaffold design for tissue engineering.

Highlights

Meshwork[18], which serves as a template for extracellular matrix (ECM) remodelling
DFs can be de-differentiated into induced pluripotent stem cells[24], and under specific culture conditions, they can be transdifferentiated into cells of other lineages, such as osteocytes[4], hepatocytes[25], macrophages[26], endothelial cells[27], and chondrocytes[3], the latter sharing many common features with NPCs28
Protein class analysis using PANTHER indicated that differentially expressed candidates in the nucleus pulposus cells (NPCs)-collagen microspheres before decellularization were mainly cytoskeletal proteins (22.6%), ECM proteins (11.3%) and nucleic acid binding (11.3%) proteins; whereas after decellularization, cytoskeletal proteins (42.9%), structural proteins (16.7%), ECM proteins (11.9%) and chaperone proteins (11.9%) were most abundant (Fig. 3B)

Summary

Introduction

Meshwork[18], which serves as a template for ECM remodelling. We have previously demonstrated that when rabbit NPCs were cultured in the collagen meshwork, they maintained their normal phenotype[19]. DFs can be de-differentiated into induced pluripotent stem cells (iPSCs)[24], and under specific culture conditions, they can be transdifferentiated into cells of other lineages, such as osteocytes[4], hepatocytes[25], macrophages[26], endothelial cells[27], and chondrocytes[3], the latter sharing many common features with NPCs28 In this project, we endeavoured to reconstitute a biomimetic NP matrix microenvironment derived from primary nucleus pulposus cells (NPCs) and analysed the composition of the NPC-derived ECM niche and investigated the effect of such niche on cultured human DFs. Our approach was to culture NPCs within collagen microspheres using methods to maintain their normal phenotype, so that they would remodel the template collagen meshwork with an NPC-derived ECM microenvironment. We believe that having a systematic understanding of the molecular composition of the cell-derived matrix and optimization of a biomimetic microenvironment is crucial for rationalized scaffold design for future tissue engineering

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