The exosome precipitation method affects the purity of the exosome and the quality of the downstream application. Polymer-based precipitation is a cost-effective method widely used in different research fields. The percentage of the polymer should be modified in different cell types or liquid biopsy before precipitation. This study aimed to optimize the protocol of the poly(ethylene glycol) (PEG)-based approach for extracting oral squamous cell carcinoma (OSCC)-derived exosomes, and its downstream applications. We used 8%, 10%, and 12% PEG to isolate the exosomes from the culture medium and compared the purity with that of the ultracentrifugation method. In addition, we extracted exosomal protein, DNA, and RNA, and tested the cell transfection efficiency for downstream application. The results reveal that 8% PEG and the medium mixture incubated at 4 °C overnight effectively precipitated exosomes of higher purity and more proper size and particle numbers compared with the ultracentrifuge method. PEG-precipitated exosomes cocultured with fibroblasts showed better transfection efficiency compared to exosomes alone. Therefore, 8% PEG is ideal for OSCC-derived exosome isolation and downstream applications. We recommend that the cost-effective PEG precipitation method be used for precipitating exosomes from OSCC cell experiments.

Dipeptidyl peptidase-4 is known to be involved in the progression of several fibrogenic diseases, but its association with oral submucous fibrosis remains unclear. This study aims to ascertain whether dipeptidyl peptidase-4 plays a role in the pathogenesis of arecoline-induced oral submucous fibrosis. We assessed the expression of dipeptidyl peptidase-4 in arecoline-treated epithelial cells and the exosomes derived from cells. We cocultured the fibroblast and exosomes derived from epithelium cells and assessed fibrogenic activity by measuring collagen secretion, α-SMA expression, and gel contraction capability. An animal study was conducted to confirm the fibrogenic activity of exosomes derived from arecoline-treated epithelial cells. Additionally, we employed a dipeptidyl peptidase-4 inhibitor to assess its efficacy in mitigating fibrogenesis. Following arecoline treatment, an increase dipeptidyl peptidase-4 expression was observed in exosomes from the treated epithelium cells. When these exosomes cocultured with fibroblast, fibrogenic gene α-SMA was upregulated, increased collagen secretion, and enhanced gel contraction capability. In a mouse model, the administration of arecoline-treated epithelium-derived exosomes induced oral submucous fibrosis phenotype, characterized by a reduction in incisal distance and epithelial atrophy. These findings offer valuable insights into clinical strategies for combating oral fibrotic disease and contribute to the foundation of future research in this field.