Presentation Information
[IO2-1]Time-Dependent Extracellular Matrix Maturation During Osteoblastic Differentiation Revealed by Integrated Transcriptomic and Proteomic Analyses
*Hlaing Pwint Phyu1, Azusa Dobashi1, Mizuki Kobayashi1, Yoshiki Ono1, Masaru Kaku1 (1. Division of Bio-Prosthodontics, Faculty of Dentistry & Graduate School of Medical and Dental Sciences, Niigata University)
[Objective]
Long-term success of prosthodontic treatments, particularly dental implant therapy, depends on proper bone quality, in which the extracellular matrix (ECM), the main organic constituent of bone, plays a central role. Although osteoblastic differentiation has been extensively investigated at the transcriptional level, how gene expression programs translate into ECM protein accumulation and maturation remains unclear. This study aimed to elucidate ECM maturation during osteoblastic differentiation by examining the temporal relationship between osteoblast gene expression and ECM deposition using an integrative transcriptomic and ECM-oriented proteomic approach.
[Method]
A mouse calvaria-derived pre-osteoblastic cell line (MC3T3-E1) was cultured for up to four weeks. Transcriptomic changes were analyzed by bulk RNA sequencing at weekly intervals. ECM composition was quantitatively analyzed using an ECM-focused proteomic workflow incorporating decellularization, hydroxylamine-based chemical digestion, deglycosylation, and LC–MS/MS analysis. Temporal changes in gene expression and ECM protein abundance were compared using integrative bioinformatic analyses.
[Results and Discussion]
Transcriptomic analysis revealed a major shift in gene expression between weeks 1 and 2, characterized by a transition from cell cycle–related pathways to osteoblast differentiation and ECM-associated pathways. In contrast, ECM proteomic analysis showed a progressive and cumulative increase in total ECM protein content throughout differentiation, despite relatively stable numbers of identified ECM proteins. Type I collagen predominated at all stages, whereas non-collagenous ECM proteins, including proteoglycans and ECM glycoproteins, exhibited distinct temporal remodeling patterns. Correlation analysis demonstrated weak concordance between mRNA expression and ECM protein abundance, indicating substantial temporal uncoupling between transcriptional regulation and matrix accumulation. Only a limited subset of proteins, such as PLOD1, PODN, and LAMB1, showed parallel transcript-level changes. These findings indicate that ECM maturation reflects cumulative osteoblastic activity rather than single-time-point gene expression. Collectively, the results highlight that bone quality is governed by time-dependent ECM maturation, providing a biological basis for understanding osseointegration, peri-implant bone stability, and long-term implant success.
Long-term success of prosthodontic treatments, particularly dental implant therapy, depends on proper bone quality, in which the extracellular matrix (ECM), the main organic constituent of bone, plays a central role. Although osteoblastic differentiation has been extensively investigated at the transcriptional level, how gene expression programs translate into ECM protein accumulation and maturation remains unclear. This study aimed to elucidate ECM maturation during osteoblastic differentiation by examining the temporal relationship between osteoblast gene expression and ECM deposition using an integrative transcriptomic and ECM-oriented proteomic approach.
[Method]
A mouse calvaria-derived pre-osteoblastic cell line (MC3T3-E1) was cultured for up to four weeks. Transcriptomic changes were analyzed by bulk RNA sequencing at weekly intervals. ECM composition was quantitatively analyzed using an ECM-focused proteomic workflow incorporating decellularization, hydroxylamine-based chemical digestion, deglycosylation, and LC–MS/MS analysis. Temporal changes in gene expression and ECM protein abundance were compared using integrative bioinformatic analyses.
[Results and Discussion]
Transcriptomic analysis revealed a major shift in gene expression between weeks 1 and 2, characterized by a transition from cell cycle–related pathways to osteoblast differentiation and ECM-associated pathways. In contrast, ECM proteomic analysis showed a progressive and cumulative increase in total ECM protein content throughout differentiation, despite relatively stable numbers of identified ECM proteins. Type I collagen predominated at all stages, whereas non-collagenous ECM proteins, including proteoglycans and ECM glycoproteins, exhibited distinct temporal remodeling patterns. Correlation analysis demonstrated weak concordance between mRNA expression and ECM protein abundance, indicating substantial temporal uncoupling between transcriptional regulation and matrix accumulation. Only a limited subset of proteins, such as PLOD1, PODN, and LAMB1, showed parallel transcript-level changes. These findings indicate that ECM maturation reflects cumulative osteoblastic activity rather than single-time-point gene expression. Collectively, the results highlight that bone quality is governed by time-dependent ECM maturation, providing a biological basis for understanding osseointegration, peri-implant bone stability, and long-term implant success.