Biochemical and Biophysical Research Communications
Mitochondrial abnormalities are involved in periodontal ligament fibroblast apoptosis induced by oxidative stress
Introduction
Periodontitis is a highly prevalent disease and affects 2.5 billion people worldwide. Moreover, it leads to the destruction of periodontal connective tissues and the loss of alveolar bone, which is regarded as the leading factor of adult tooth loss [1,2]. PDL cells (PDLCs), the major cell type of periodontal ligament (PDL), are considered important participants of periodontal metabolism and remodeling [3]. The number of PDLCs is kept relatively stable through the balance of cell proliferation and death under physiological conditions [4]. Various pathological factors, including periodontal pathogens, cytokines, and drugs can lead to PDLCs apoptosis, accounted for periodontitis [5,6]. However, the mechanisms underlying apoptosis of PDLCs are not fully understood. Perhaps this is why the therapies available for treating periodontitis are not very efficacious. Therefore, the mechanisms related to PDLCs apoptosis must be further explored to identify effective therapeutic approaches to periodontitis.
Accumulating evidence suggests that oxidative stress (OS) features overproduction of highly reactive oxygen species (ROS) owing to the disturbed pro-oxidant/antioxidant balance, playing a key role in the onset and progression of inflammation [7]. Moreover, studies indicate that periodontal-tissue destruction closely correlates with excessive ROS production and with compromised antioxidant capacity in periodontitis [8]. Notably, administration of antioxidants can effectively inhibit the development of periodontitis, partially through alleviation of PDLCs apoptosis induced by OS [9]. Thus, investigating the mechanisms by which OS affects the apoptosis of PDLCs represents one of the primary tasks in the search for effective treatment of periodontitis.
Mitochondria are the primary source of cellular ROS and are subject to a direct attack by ROS [10]. Overproduced ROS cause oxidative damage to mitochondrial proteins and structures, thus subsequently leading to mitochondrial dysfunction [11]. The latter manifests itself as decreases in mitochondrial membrane potential and in uncoupled electron transport (in the electron transport chain) along with ATP downregulation, which affects the function of cells and subsequently results in apoptosis [12]. Our previous study has revealed that mitochondrial dysfunction is closely associated with aggravation of periodontitis caused by diabetes [13]. In addition, mitochondrial dysfunction is considered an important contributing factor of apoptosis of osteoblasts and of Hep-G2 cells under the influence of OS [14,15]. However, few studies have comprehensively examined the participation of mitochondrial dysfunction in periodontal-cell apoptosis caused by OS.
Furthermore, mitochondria are organelles that dynamically alter their shape by frequent fusion and fission to continuously perform their function in the cell. Mitochondrial dynamics are not only critical for the homeostasis of mitochondria but also important for the well-being of the whole cell [16]. ROS are important for regulating the mitochondrial dynamics during several physiological and pathological processes [17]. Moreover, it is known that the loss of mitochondrial function, secondary to defects in mitochondrial dynamics, also leads to an increase of ROS generation [18]. These studies indicate that impaired mitochondrial dynamic leads to mitochondrial dysfunction and OS, which induces cellular perturbation and injury. It is still unclear, however, whether this mitochondrial fusion-and-fission machinery is active in OS-affected PDLCs. Therefore, in this study, our objectives were to investigate (1) whether mitochondrial dysfunction participates in the OS-induced apoptosis of human PDLCs (hPDLCs) and (2) whether mitochondrial dynamics act as a regulator of hPDLCs apoptosis induced by OS. We found that improvement in mitochondrial function exerted a significant protective effect against the hPDLCs apoptosis induced by OS. Thus, this result may have significant implications for therapeutic interventions in periodontitis.
Section snippets
Cell culture and treatment
The study protocol was approved by the Committee on Research on Human Subjects of the School and Hospital of Stomatology, Wenzhou Medical University. hPDLCs were obtained from healthy third molars of five healthy men aged 20–30 years (mean age: 24.6) who underwent molar extraction after providing informed consent. PDL tissues were extracted from the root surface of a tooth and were cut into 1–2 mm2 pieces. The tissue was cultured in α-minimum essential medium containing 10% of fetal bovine
H2O2-induced hPDLCs apoptosis
The MTT results showed that after H2O2 treatment, cell viability significantly decreased in a time- and dose-dependent manner (Fig. 1A). The results of flow cytometry indicated that the apoptosis rate increased with treatment duration. Early apoptosis was significant after 0.3 mM H2O2 treatment for 12 h, and 0.3 mM H2O2 for 24 h induced late apoptosis and slightly increased necrosis (Fig. 1B–C). H2O2-induced DNA damage was next confirmed by obviously increased TUNEL-positive staining (Fig. 1
Discussion
Although OS-induced PDLCs apoptosis plays a critical role in the initiation and progression of periodontitis, the mechanism behind this phenomenon is still unclear. Emerging evidence indicates that mitochondrial dysfunction is closely linked to OS. However, whether mitochondrial dysfunction is involved in PDLCs apoptosis induced by OS is poorly understood. In the current study, we clearly demonstrated that mitochondrial dysfunction and impaired mitochondrial dynamics were evident in the process
Conflicts of interest
All authors declare that they have no competing interests.
Acknowledgments
This work was supported by the following grants: the Natural Science Foundation of China (No. 81500817, 81870777, and 81802230), Zhejiang Provincial Natural Science Foundation of China (No. LY15H140008), Health Science and Technology Project of Zhejiang Province (No. 2016KYB184), Zhejiang Provincial Science and Technology Project for Public Welfare (No. 2017C33081), Wenzhou Municipal Science and Technology Project for Public Welfare (No. Y20150069 and Y20170026), Wenzhou Science and Technology
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