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The activity, distribution, and colocalization of cathepsin K and matrix metalloproteases in intact and eroded dentin

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Abstract

Objectives

This study aimed to assess the activity, distribution, and colocalization of cathepsin K (catK) and matrix metalloproteases (MMPs) in both intact and eroded dentin in vitro.

Materials and methods

Eroded dentin was obtained by consecutive treatment with 5% citric acid (pH = 2.3) for 7 days, while intact dentin remained untreated. Pulverized dentin powder (1.0 g) was extracted from both intact and eroded dentin using 5 mL of 50 mM Tris–HCl buffer (0.2 g/1 mL, pH = 7.4) for 60 h to measure the activity of catK and MMPs spectrofluorometrically. In addition, three 200-μm-thick dentin slices were prepared from intact and eroded dentin for double-labeling immunofluorescence to evaluate the distribution and colocalization of catK and MMPs (MMP-2 and MMP-9). The distribution and colocalization of enzymes were analyzed using inverted confocal laser scanning microscopy (CLSM), with colocalization rates quantified using Leica Application Suite Advanced Fluorescent (LAS AF) software. One-way analysis of variance (ANOVA) was used to analyze the fluorescence data related to enzyme activity (α = 0.05).

Results

The activity of catK and MMPs was significantly increased in eroded dentin compared with intact dentin. After erosive attacks, catK, MMP-2, and MMP-9 were prominently localized in the eroded regions. The colocalization rates of catK with MMP-2 and MMP-9 were 13- and 26-fold higher in eroded dentin, respectively, than in intact dentin.

Conclusions

Erosive attacks amplified the activity of catK and MMPs in dentin while also altering their distribution patterns. Colocalization between catK and MMPs increased following erosive attacks.

Clinical relevance

CatK, MMP-2, and MMP-9 likely play synergistic roles in the pathophysiology of dentin erosion.

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Data availability

The data that support the findings of this study are available from the corresponding author upon reasonable request.

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Acknowledgements

The authors would like to thank Ling Lin from the Public Technology Service Center of Fujian Medical University for technical assistance.

Funding

This research project was supported by the Fujian Provincial Health Technology Project (2022CXA042) and the Fujian Provincial Special Financial Funds (2023CZZX01).

Author information

Author notes

  1. Xiujiao Lin and Xinwen Tong contributed equally to this work.

Authors and Affiliations

  1. Fujian Key Laboratory of Oral Diseases & Fujian Provincial Engineering Research Center of Oral Biomaterial & Stomatological Key Laboratory of Fujian College and University, School and Hospital of Stomatology, Fujian Medical University, Fuzhou, China

    Xiujiao Lin, Xinwen Tong, Yiying Chen & Hao Yu

  2. Research Center of Dental Esthetics and Biomechanics, Fujian Medical University, Fuzhou, China

    Xiujiao Lin, Xinwen Tong, Yiying Chen & Hao Yu

  3. Department of Stomatology, Zhangzhou Affiliated Hospital of Fujian Medical University, Zhangzhou, China

    Xiujiao Lin

  4. Public Technology Service Center, Fujian Medical University, Fuzhou, China

    Zhihong Huang

  5. Department of Applied Prosthodontics, Graduate School of Biomedical Sciences, Nagasaki University, Nagasaki, Japan

    Hao Yu

  6. Department of Prosthodontics, School and Hospital of Stomatology, Fujian Medical University, Yangqiao Zhong Road 246, Fuzhou, 350000, China

    Hao Yu

Authors
  1. Xiujiao Lin
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Contributions

Xiu-jiao Lin and Xin-wen Tong wrote the original draft and performed the data curation. Yi-ying Chen performed the formal analysis and the investigation. Zhi-hong Huang prepared the materials and performed the investigation. Hao Yu conceptualized and supervised the study and reviewed and edited the manuscript. All authors have read and approved the final manuscript.

Corresponding author

Correspondence to Hao Yu.

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Ethical approval

All procedures performed in the present study were in accordance with the ethical standards of the institutional research committee and with the 1964 Helsinki Declaration and its later amendments or comparable ethical standards. Ethical approval for the study was granted by the Ethics Committee of the School and Hospital of Stomatology, Fujian Medical University (Approval no. 2022CXA042).

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Lin, X., Tong, X., Chen, Y. et al. The activity, distribution, and colocalization of cathepsin K and matrix metalloproteases in intact and eroded dentin. Clin Oral Invest 28, 1 (2024). https://doi.org/10.1007/s00784-023-05393-5

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