ERS Mediated by GRP-78/PERK/CHOP Signaling Is Involved in Fluoride-Induced Ameloblast Apoptosis.

Fluoride can be widely ingested from the environment, and its excessive intake could result in adverse effects. Dental fluorosis is an early sign of fluoride toxicity which can cause esthetic and functional problems. Though apoptosis in ameloblasts is one of the potential mechanisms, the specific signal cascade is in-conclusive. High-throughput sequencing and molecular biological techniques were used in this study to explore the underlying pathogenesis of dental fluorosis, for its prevention and treatment. A fluorosis cell model was established. Viability and apoptosis rate of mouse ameloblast-derived cell line (LS8 cells) was measured using cell counting kit-8 (CCK-8) assay and flow cytometry analysis. Cells were harvested with or without 2-mM sodium fluoride (NaF) stimulation for high-throughput sequencing. Based on the sequencing data, subcellular structures, endoplasmic reticulum stress (ERS), and apoptosis related biomarkers were verified using transmission electron microscopy, quantitative real-time polymerase chain reaction, and Western blotting techniques. Expression of ERS markers, apoptosis related proteins, and enamel formation enzymes were detected using Western blotting after addition of 4-phenylbutyrate (4-PBA). NaF-inhibited LS8 cells displayed time- and dose- dependent viability. Additionally, apoptosis and morphological changes were observed. RNA-sequencing data showed that protein processing in endoplasmic reticulum was obviously affected. ERS and apoptosis were induced by excessive NaF. Downregulation of kallikrein-related peptidase 4 (KLK4) was also observed. Inhibition of ERS by 4-PBA rescued the apoptotic and functional protein changes in cells. Excessive fluoride induces apoptosis by activating ERS, which is mediated by GRP-78/PERK/CHOP signaling. Key proteinase is present in maturation-stage enamel; KLK4 was also affected by fluoride, but rescued by 4-PBA. This study presents a possibility for therapeutic strategies for dental fluorosis, while further exploration is required.

[Study on using laser Doppler flowmetry to determine the pulpal blood flow of immature maxillary incisors].

OBJECTIVE: This study aimed to evaluate the influence of age on the pulpal blood flow (PBF) of immature maxillary incisors of maxillary incisors, which was detected by laser Doppler flowmetry (LDF). METHODS: LDF was used to detect the PBF value of maxillary central and lateral incisors of a child group (aged 7-13 years old) and a positive control group (aged 18-25 years old), as well as the central incisor of a negative control group (the central incisor had undergone endodontic treatment). We then compared the features of PBF in all groups with the influence of gender and position on PBF. The relation of maxillary central incisor and lateral incisor, age, and maxillary incisor were analyzed. RESULTS: The PBF value of the negative control group was (2.08±0.73) PU. The PBF values in the positive control group in central and lateral incisors were (8.49±1.88) and (7.52±1.82) PU. In the child group, PBF values in central incisors and lateral incisors were (11.31±2.21) and (12.18±2.65) PU. A significant difference was observed between different groups and between central and lateral incisors (P<0.01). Meanwhile, no significant difference was found in the PBF values between the right and the left parts in both males and females (P>0.05). Age had a linearity negative correlation with the PBF value of incisors in the child group. A linear negative correlation existed between the age and PBF of central and lateral incisors (r=-0.310 and r=-0.510, respectively) (P<
0.01). CONCLUSIONS: PBF value decreased with increased age in children aged 7-13 years old.