Identification of exosomal circSLC26A4 as a liquid biopsy marker for cervical cancer

Abstract

Objective: Circular RNA SLC26A4 (circSLC26A4) functions as an oncogene in the initiation and progression of cervical cancer (CC). However, the clinical role of plasma exosomal circSLC26A4 in CC is poorly known. This study aims to develop an accurate diagnostic method based on circulating exosomal circSLC26A4.

Methods: In this study, exosomal circSLC26A4 derived from CC cell lines (CaSki, SiHa, and HeLa) and human cervical epithelial cells (HcerEpic) was measured and compared using quantitative reverse transcriptase polymerase chain reaction (qRT-PCR). Additionally, 56 volunteers, including 18 CC patients, 18 cervical high-grade squamous intraepithelial lesion (HSIL) patients, and 20 healthy volunteers, were enrolled. qRT-PCR was also performed to measure the plasma exosomal circSLC26A4 levels in all participants.

Results: The exosomal circSLC26A4 expression level derived from CC cells was significantly elevated compared to it derived from HcerEpic cells. Plasma exosomal circSLC26A4 levels in CC patients were significantly higher than in healthy women and HSIL patients (P < 0.05). In addition, high plasma exosomal circSLC26A4 expression was positively associated with lymph node metastasis and FIGO stage (all P < 0.05). However, no significant correlation was found between plasma exosomal circSLC26A4 expression and age, intravascular cancerous embolus, and perineural invasion (P > 0.05).

Conclusions: The high exosomal circSLC26A4 expression is closely related to the occurrence of CC. Plasma exosomal circSLC26A4 can be used as a diagnostic marker for CC.

Fig 1. The circSLC26A4 is overexpressed in the exosome of CC cell lines culture medium.

(A) Representative transmission electron microscopy (TEM) images of cell culture medium exosome. Scale bar, 100 nm. (B) Particle size distribution of cell culture medium exosome was measured using nanoparticle tracking analysis (NTA). (C) Protein immunoblots of exosomes, including the typical markers (flotillin 2, CD63). (D) RT-PCR assay was performed to measure the circSLC26A4 expression in different CC cell lines (SiHa, HeLa, and CaSki) and HcerEpic cell line medium-derived exosomes were described as a control. n = 12.

Fig 2. The circSLC26A4 is overexpressed in the plasma exosome of CC patients.

(A) Representative transmission electron microscopy (TEM) images of plasma exosome. Scale bar, 100 nm. (B) Particle size distribution of plasma exosome was measured using nanoparticle tracking analysis (NTA). (C) Protein immunoblots of exosomes, including the typical markers (flotillin 2, CD63). (D) RT-PCR indicated the circSLC26A4 expression in CC and healthy plasma.

Fig 3. Diagnostic value of ROC curve analysis of plasma exosomal circSLC26A4.

Diagnostic value of the plasma exosomal circSLC26A4 in differentiating CC patients from controls (A), CC and HSIL patients (B), and HSIL patients and controls(C).