4.7 Article

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

Journal

Publisher

MDPI
DOI: 10.3390/ijms21155273

Keywords

salivary small extracellular vesicles; ultracentrifuge; size exclusion chromatography; DNA methylation; cellular uptake

Funding

  1. University of Queensland Early Career Research Grant [UQECR194615]
  2. UQ Development Fellowship [UQFEL1716821]
  3. Lions Medical Research Foundation
  4. National Health and Medical Research Council [1114013]
  5. Fondo Nacional de Desarrollo Cientifico y Tecnologico [1170809]
  6. National Health and Medical Research Council of Australia [1114013] Funding Source: NHMRC

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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)-alpha, IL-1 beta, 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 (n= 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.

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