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Proteomic analysis of human periodontal ligament cells under hypoxia.

ABSTRACT: Background:The periodontal ligament is essential for homeostasis of periodontal tissue. A hypoxic milieu of the periodontal tissue is generated under periodontitis or during orthodontic treatment, which affects the periodontal and bone remodelling process. Here, we provide a comprehensive proteomic characterization of periodontal ligament cells under hypoxic conditions, aiming to reveal previously unappreciated biological changes and to help advance hypoxia-based therapeutic strategies for periodontal diseases. Methods:Human periodontal ligament cells (hPDLCs) were characterized using immunohistochemistry (IHC) and flow cytometry (FACS). Successful hypoxia treatment of hPDLCs with 1% O2 was confirmed by qRT-PCR. Proliferation was evaluated using an MTT assay. The proteomic expression profile under hypoxia was studied with the isobaric tags for relative and absolute quantification (iTRAQ) approach followed by protein identification and bioinformatic analysis, and western blot verification was performed. Results:The hPDLCs were positive for vimentin, CD73 and CD105 and negative for keratin, CD34 and CD45. After hypoxia treatment, the mRNA expression of hypoxia-inducible factor 1a (HIF1a) was upregulated. The proliferation rate was elevated during the first 6?h but decreased from 6?h to 72?h. A total of 220 differentially expressed proteins were quantified in hPDLCs under hypoxia (1% O2, 24?h), including 153 upregulated and 67 downregulated proteins, five of which were verified by western blot analysis. The Gene Ontology enriched terms included the energy metabolic process, membrane-bound organelle and vesicle, and protein binding terms. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated several involved pathways, including glycolysis/gluconeogenesis, biosynthesis of amino acids, the HIF-1 signalling pathway, and focal adhesion. A protein-protein interaction (PPI) network demonstrated the dominant role of autophagy over apoptosis under hypoxia. Conclusion:The proteomic profile of hPDLCs under hypoxia was mainly related to energy metabolism, autophagy, and responses to stimuli such as adhesion and inflammation. Previously unrecognized proteins including solute carrier family proteins, heat shock proteins, ubiquitination-related enzymes, collagen and S100 family proteins are involved in adaptive response to hypoxia in hPDLCs and are thus of great research interest in future work.

SUBMITTER: Li Q

PROVIDER: S-EPMC6717648 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Proteomic analysis of human periodontal ligament cells under hypoxia.

Li Qiwen Q Luo Tao T Lu Wenxin W Yi Xiaoxiao X Zhao Zhihe Z Liu Jun J

Proteome science 20190831

<h4>Background</h4>The periodontal ligament is essential for homeostasis of periodontal tissue. A hypoxic milieu of the periodontal tissue is generated under periodontitis or during orthodontic treatment, which affects the periodontal and bone remodelling process. Here, we provide a comprehensive proteomic characterization of periodontal ligament cells under hypoxic conditions, aiming to reveal previously unappreciated biological changes and to help advance hypoxia-based therapeutic strategies f ...[more]

PMID: 31496921

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Project description:The periodontal ligament (PDL) is a specialized connective tissue that provides structural support to the tooth and is crucial for oral functions. The mechanical properties of the PDL are mainly derived from the tissue-specific composition and structural characteristics of the extracellular matrix (ECM). The ECM also plays key roles in determining cell fate in the cellular microenvironment thus crucial in the PDL tissue homeostasis. In the present study, we determined the comprehensive ECM profile of mouse molar PDL using laser microdissection and mass spectrometry-based proteomic analysis with ECM-oriented data curation. Additionally, we evaluated changes in the ECM proteome under mechanical loading using a mouse orthodontic tooth movement (OTM) model and analyzed potential regulatory networks using a bioinformatics approach. Proteomic changes were evaluated in reference to the novel second harmonic generation (SHG)-based fiber characterization. Our ECM-oriented proteomics approach succeeded in illustrating the comprehensive ECM profile of the mouse molar PDL. We revealed the presence of type II collagen in PDL, possibly associated with the load-bearing function upon occlusal force. Mechanical loading induced unique architectural changes in collagen fibers along with dynamic compositional changes in the matrisome profile, particularly involving ECM glycoproteins and matrisome-associated proteins. We identified several unique matrisome proteins which responded to the different modes of mechanical loading in PDL. Notably, the proportion of type VI collagen significantly increased at the mesial side, contributing to collagen fibrogenesis. On the other hand, type XII collagen increased at the PDL-cementum boundary of the distal side. Furthermore, a multifaceted bioinformatics approach illustrated the potential molecular cues, including PDGF signaling, that maintain ECM homeostasis under mechanical loading. Our findings provide fundamental insights into the molecular network underlying ECM homeostasis in PDL, which is vital for clinical diagnosis and development of biomimetic tissue-regeneration strategies.

Dataset Information

Proteomic analysis of human periodontal ligament cells under hypoxia.

Publications

Proteomic analysis of human periodontal ligament cells under hypoxia.

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