褪黑素通过抑制 MMP3 衰减体外扩增的牙髓干细胞衰老。

Melatonin attenuates dental pulp stem cells senescence due to vitro expansion via inhibiting MMP3.

机构信息

Department of Endodontics, Liaoning Provincial Key Laboratory of Oral Diseases, School and Hospital of Stomatology, China Medical University, Shenyang, China.

Department of Cardiology, The First Affiliated Hospital of China Medical University, Shenyang, Liaoning, China.

出版信息

Oral Dis. 2024 May;30(4):2410-2424. doi: 10.1111/odi.14649. Epub 2023 Jul 14.

DOI:10.1111/odi.14649

PMID:37448325

Abstract

OBJECTIVE

We aimed to identify the crucial genes involved in dental pulp stem cell (DPSC) senescence and evaluate the impact of melatonin on DPSC senescence.

METHODS

Western blotting, SA-β-Gal staining and ALP staining were used to evaluate the senescence and differentiation potential of DPSCs. The optimal concentration of melatonin was determined using the CCK-8 assay. Differentially expressed genes (DEGs) involved in DPSC senescence were obtained via bioinformatics analysis, followed by RT-qPCR. Gain- and loss-of-function studies were conducted to explore the role of MMP3 in DPSC in vitro expansion and in response to melatonin. GSEA was employed to analyse MMP3-related pathways in cellular senescence.

RESULTS

Treatment with 0.1 μM melatonin attenuated cellular senescence and differentiation potential suppression in DPSCs due to long-term in vitro expansion. MMP3 was a crucial gene in senescence, as confirmed by bioinformatics analysis, RT-qPCR and Western blotting. Furthermore, gain- and loss-of-function studies revealed that MMP3 played a regulatory role in cellular senescence. Rescue assays showed that overexpression of MMP3 reversed the effect of melatonin on senescence. GSEA revealed that the MMP3-dependent anti-senescence effect of melatonin was associated with the IL6-JAK-STAT3, TNF-α-Signalling-VIA-NF-κB, COMPLEMENT, NOTCH Signalling and PI3K-AKT-mTOR pathways.

CONCLUSION

Melatonin attenuated DPSC senescence caused by long-term expansion by inhibiting MMP3.

摘要

目的

本研究旨在鉴定与牙髓干细胞（DPSC）衰老相关的关键基因，并评估褪黑素对 DPSC 衰老的影响。

方法

采用 Western blot、SA-β-Gal 染色和 ALP 染色评估 DPSCs 的衰老和分化潜能。使用 CCK-8 法确定褪黑素的最佳浓度。通过生物信息学分析获得与 DPSC 衰老相关的差异表达基因（DEGs），并进行 RT-qPCR。进行 gain-和 loss-of-function 研究，以探讨 MMP3 在 DPSC 体外扩增和对褪黑素反应中的作用。采用 GSEA 分析 MMP3 相关通路在细胞衰老中的作用。

结果

0.1μM 褪黑素处理可减轻由于长期体外扩增导致的 DPSCs 细胞衰老和分化潜能抑制。生物信息学分析、RT-qPCR 和 Western blot 证实 MMP3 是衰老的关键基因。此外，gain-和 loss-of-function 研究表明 MMP3 在细胞衰老中起调节作用。挽救实验表明，MMP3 的过表达逆转了褪黑素对衰老的影响。GSEA 显示，褪黑素通过 MMP3 发挥抗衰老作用与 IL6-JAK-STAT3、TNF-α-Signalling-VIA-NF-κB、COMPLEMENT、NOTCH Signalling 和 PI3K-AKT-mTOR 通路有关。