Increased apoptosis of gingival epithelium is associated with impaired autophagic flux in medication-related osteonecrosis of the jaw

ABSTRACT

Macroautophagy/autophagy has both negative and positive aspects in the development of many diseases. Yet, its exact role and specific mechanism in the onset of medication-related osteonecrosis of the jaw (MRONJ) is still not fully understood. Retarded gingiva healing is the primary clinical manifestation in patients with MRONJ. In this study, we aimed to explore the relationship between autophagy and apoptosis in MRONJ gingival epithelium and search for a method to prevent this disease. First, we examined clinical samples from patients diagnosed with MRONJ and healthy controls, finding that autophagy-related markers MAP1LC3/LC3 and SQSTM1/p62 synchronously increased, thus suggesting that autophagic flux was suppressed in MRONJ. Moreover, mRNA sequencing analysis and TUNEL assay showed that the process of apoptosis was upregulated in patients and animals with MRONJ, indicating autophagy and apoptosis participate in the development of MRONJ. Furthermore, the level of autophagy and apoptosis in zoledronic acid (ZA)-treated human keratinocytes cell lines (HaCaT cells) was concentration dependent in vitro. In addition, we also found that RAB7 (RAB7, member RAS oncogene family) activator ML098 could rescue MRONJ gingival lesions in mice by activating the autophagic flux and downregulating apoptosis. To sum up, this study demonstrated that autophagic flux is impaired in the gingival epithelium during MRONJ, and the rescued autophagic flux could prevent the occurrence of MRONJ.

Abbreviations: ACTB: actin beta; Baf-A1: bafilomycin A₁; CASP3: caspase 3; CASP8: caspase 8; CT: computed tomography; DMSO: dimethyl sulfoxide; GFP: green fluorescent protein; HaCaT cells: human keratinocytes cell lines; H&E: hematoxylin and eosin; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; MRONJ: medication-related osteonecrosis of the jaw; PARP: poly(ADP-ribose) polymerase; RAB7: RAB7, member RAS oncogene family; RFP: red fluorescent protein; SQSTM1/p62: sequestosome 1; TEM: transmission electron microscopy; ZA: zoledronic acid.

KEYWORDS:

• Apoptosis
• autophagic flux
• gingival epithelium
• keratinocytes
• medication-related osteonecrosis of the jaw
• zoledronic acid

Acknowledgements

We wish to thank the Central laboratory of Peking University School and Hospital of Stomatology for providing the research facilities. We thank Pei Fei for the valuable discussion. We also thank Cloud-Seq Biotech Ltd. Co. (Shanghai, China) for the RNA high-throughput sequencing service and the subsequent analysis.

Disclosure statement

No potential conflict of interest was reported by the authors.

Data availability statement

The authors confirm that the data supporting the findings of this study are available within the article [and/or] its supplementary materials.

Supplementary material

Supplemental data for this article can be accessed online at https://doi.org/10.1080/15548627.2023.2234228

Additional information

Funding

The work was supported by the National Natural Science Foundation of China [82101041]; Beijing Natural Science Foundation [7202234]; Beijing Natural Science Foundation [7222223]; the Youth Found of Peking University School and Hospital of Stomatology [PKUSS20200110].