Detection of Salivary Small Extracellular Vesicles Associated Inflammatory Cytokines Gene Methylation in Gingivitis

• Main Authors: Pingping Han, Andrew Lai, Carlos Salomon, Sašo Ivanovski
• Format: Article
• Language: English
• Published: MDPI AG 2020-07-01
• Series: International Journal of Molecular Sciences
• Subjects: salivary small extracellular vesicles; ultracentrifuge; size exclusion chromatography; DNA methylation; cellular uptake
• Online Access: https://www.mdpi.com/1422-0067/21/15/5273

Salivary small extracellular vesicles (sEV) are emerging as a potential liquid biopsy for oral diseases. However, technical difficulties for salivary sEV isolation remain a challenge. Twelve participants (five periodontally healthy, seven gingivitis patients) were recruited and salivary sEV were isolated by ultracentrifuge (UC-sEV) and size exclusion chromatography (SEC-sEV). The effect of UC and SEC on sEV yield, DNA methylation of five cytokine gene promoters (interleukin (IL)−6, tumor necrosis factor (TNF)-α, IL−1β, IL−8, and IL−10), and functional uptake by human primary gingival fibroblasts (hGFs) was investigated. The results demonstrated that SEC-sEV had a higher yield of particles and particle/protein ratios compared to UC-sEV, with a minimal effect on the detection of DNA methylation of five cytokine genes and functional uptake in hGFs (<i>n</i> = 3). Comparing salivary sEV characteristics between gingivitis and healthy patients, gingivitis-UC-sEV were increased compared to the healthy group; while no differences were found in sEV size, oral bacterial gDNA, and DNA methylation for five cytokine gene promoters, for both UC-sEV and SEC-sEV. Overall, the data indicate that SEC results in a higher yield of salivary sEV, with no significant differences in sEV DNA epigenetics, compared to UC.