Pediatric Surgery, Children's Hospital of Changchun, Changchun, 130021, Jilin Province, People's Republic of China.
Department of Stomatology, The First Hospital of Jilin University, Changchun, 130021, Jilin Province, People's Republic of China.
Odontology. 2023 Oct;111(4):870-882. doi: 10.1007/s10266-023-00793-1. Epub 2023 Mar 6.
Periodontal tissue regeneration engineering based on human periodontal ligament stem cells (hPDLSCs) provides a broad prospect for the treatment of periodontal disease. N-Acetyltransferase 10 (NAT10)-catalyzed non-histone acetylation is widely involved in physiological or pathophysiological processes. However, its function in hPDLSCs is still missing. hPDLSCs were isolated, purified, and cultured from extracted teeth. Surface markers were detected by flow cytometry. Osteogenic, adipogenic, and chondrogenic differentiation potential was detected by alizarin red staining (ARS), oil red O staining, and Alcian blue staining. Alkaline phosphatase (ALP) activity was assessed by ALP assay. Quantitative real-time PCR (qRT-PCR) and western blot were used to detect the expression of key molecules, such as NAT10, Vascular endothelial growth factor A (VEGFA), PI3K/AKT pathway, as well as bone markers (RUNX2, OCN, OPN). RNA-Binding Protein Immunoprecipitation PCR (RIP-PCR) was used to detect the N4-acetylcytidine (ac4C) mRNA level. Genes related to VEGFA were identified by bioinformatics analysis. NAT10 was highly expressed in the osteogenic differentiation process with enhanced ALP activity and osteogenic capability, and elevated expression of osteogenesis-related markers. The ac4C level and expression of VEGFA were obviously regulated by NAT10 and overexpression of VEGFA also had similar effects to NAT10. The phosphorylation level of PI3K and AKT was also elevated by overexpression of VEGFA. VEGFA could reverse the effects of NAT10 in hPDLSCs. NAT10 enhances the osteogenic development of hPDLSCs via regulation of the VEGFA-mediated PI3K/AKT signaling pathway by ac4C alteration.
基于人牙周膜干细胞（hPDLSCs）的牙周组织再生工程为牙周病的治疗提供了广阔的前景。N-乙酰基转移酶 10（NAT10）催化的非组蛋白乙酰化广泛参与生理或病理生理过程。然而，其在 hPDLSCs 中的功能尚不清楚。从提取的牙齿中分离、纯化和培养 hPDLSCs。通过流式细胞术检测表面标志物。通过茜素红染色（ARS）、油红 O 染色和阿利新蓝染色检测成骨、成脂和成软骨分化潜能。通过碱性磷酸酶（ALP）测定法评估 ALP 活性。通过定量实时 PCR（qRT-PCR）和 Western blot 检测关键分子（如 NAT10、血管内皮生长因子 A（VEGFA）、PI3K/AKT 通路以及骨标志物（RUNX2、OCN、OPN）的表达。RNA 结合蛋白免疫沉淀 PCR（RIP-PCR）用于检测 N4-乙酰胞苷（ac4C）mRNA 水平。通过生物信息学分析鉴定与 VEGFA 相关的基因。NAT10 在成骨分化过程中高表达，具有增强的 ALP 活性和成骨能力，以及升高的成骨相关标志物表达。ac4C 水平和 VEGFA 的表达受 NAT10 调节，而过表达 VEGFA 也具有类似于 NAT10 的作用。VEGFA 还能上调 PI3K 和 AKT 的磷酸化水平。VEGFA 可以逆转 NAT10 在 hPDLSCs 中的作用。NAT10 通过 ac4C 改变调节 VEGFA 介导的 PI3K/AKT 信号通路增强 hPDLSCs 的成骨发育。
