Huang Xin, Kuang Shuhong, Shen Zongshan, Liang Min, Lin Zhengmei
Department of Periodontology, Guanghua School and Hospital of Stomatology, Sun Yat-Sen University, Guangzhou, Guangdong, China.
Guangdong Provincial Key Laboratory of Stomatology, Sun Yat-Sen University, North Campus, Guangzhou, Guangdong, China.
J Periodontol. 2020 May;91(5):705-714. doi: 10.1002/JPER.19-0262. Epub 2019 Oct 24.
Hyperglycemic micro-environment induced by diabetes could regulate the response of periodontal tissues to pathogenic microorganisms in which disruption of autophagy lysosomal pathway (ALP) may participate. This study aimed to explore the mechanisms underlying how high glucose (HG) regulates ALP in gingival epithelial cells (GECs).
Human gingival tissues in healthy group (C), periodontitis group (P), diabetes group (DM), and diabetes + periodontitis group (DP) were collected and were applied to observe the fusion of autophagy and lysosome. Diabetic mouse model with periodontitis was established and RNA-seq was applied to investigate the expression of ALP-associated genes in gingival epithelium. To explore the key role of ATPase transmembrane v0 domain, subunit c (ATP6V0C) in the disruption of ALP by HG, human gingival epithelial cells (HGECs) were cultured in 5.5 mM/25 mM glucose medium for 48 hours and followed by Porphyromonas gingivalis stimulation for 0, 6, and 12 hours. HBLV-h-ATP6V0C was transfected in HGECs that were stimulated by 25 mM HG condition.
Immunofluorescence double staining exhibited the disruption of ALP in human gingival epithelium in diabetes groups and HGECs under 25 mM glucose condition, accompanied with significantly downregulated lysosomal acidity. RNA-seq of mouse gingival epithelium screened out Atp6v0c. Compared with HGECs in normal culture medium, ATP6V0C expression and LC3-II/I expression ratio were significantly downregulated, with an upregulated expression of P62, IL-1β in HGECs under HG condition. Over-expression of ATP6V0C rescued HG-induced disruption of ALP in HBLV-h-ATP6V0C transfected HGECs, with significantly upregulation of LC3-II/I and downregulation of P62, IL-1β.
ATP6V0C mediates HG-induced ALP disruption in HGECs, eventually exacerbates periodontal inflammation.
糖尿病诱导的高血糖微环境可调节牙周组织对致病微生物的反应，其中自噬溶酶体途径（ALP）的破坏可能参与其中。本研究旨在探讨高糖（HG）调节牙龈上皮细胞（GECs）中ALP的潜在机制。
收集健康组（C）、牙周炎组（P）、糖尿病组（DM）和糖尿病合并牙周炎组（DP）的人牙龈组织，用于观察自噬与溶酶体的融合。建立糖尿病伴牙周炎小鼠模型，并应用RNA测序研究牙龈上皮中ALP相关基因的表达。为了探讨ATP酶跨膜v0结构域亚基c（ATP6V0C）在HG破坏ALP中的关键作用，将人牙龈上皮细胞（HGECs）在5.5 mM/25 mM葡萄糖培养基中培养48小时，然后用牙龈卟啉单胞菌刺激0、6和12小时。将HBLV-h-ATP6V0C转染到受25 mM HG条件刺激的HGECs中。
免疫荧光双重染色显示糖尿病组人牙龈上皮和25 mM葡萄糖条件下的HGECs中ALP被破坏，同时溶酶体酸度显著下调。小鼠牙龈上皮的RNA测序筛选出Atp6v0c。与正常培养基中的HGECs相比，HG条件下HGECs中ATP6V0C表达和LC3-II/I表达比值显著下调，P62、IL-1β表达上调。ATP6V0C的过表达挽救了HBLV-h-ATP6V0C转染的HGECs中HG诱导的ALP破坏，LC3-II/I显著上调，P62、IL-1β下调。
ATP6V0C介导HG诱导的HGECs中ALP破坏，最终加剧牙周炎症。
