Human papillomavirus E6E7 mRNA and TERC lncRNA in situ detection in cervical scraped cells and cervical disease progression assessment

BACKGROUND: Human papillomavirus screen in female cervical cells has demonstrated values in clinical diagnosis of precancerous lesions and cervical cancers. Human papillomavirus tests of cervical cells by utilizing Polymerase Chain Reaction (PCR) method provides human papillomavirus infection status however no further virus in situ information. Although it is well known that the tests of human papillomavirus E6/E7 RNA location in infected cervical cells and cell internal malignancy molecular will provide clues for gynecologists to evaluate disease progression, there are technique difficulties to preserve RNAs in cervical scraped cells for in situ hybridization. METHODS: In current study, after developing a cervical cell collection and preparation method for RNA in situ hybridization, we captured the chance to screen 98 patient cervical cell samples and detected human papillomavirus E6/E7 mRNAs of high-risk subtypes, low-risk subtypes and long non-coding RNA (lncRNA) TERC in the cells. RESULTS: There were 69 samples exhibited consistence between human papillomavirus PCR and human papillomavirus RNA in situ hybridization results in cervical collected cells. Among them, 23 were both positive and 46 were both negative. In the rest 29 samples, 8 were HPV RNAscope positive, either high risk or low risk subtypes, however HPV PCR negative. Another 9 samples were HPV PCR results positive whereas RNAscope negative. The last 12 samples were HPV positive detected by both RNAscope and PCR methods, however inconsistent between high-risk and low-risk subtypes. In RNAscope positive samples, viral E6/E7 mRNAs were observed to distribute in cervical scraped cell nucleus and cytoplasm. Moreover, HPV viral RNA gathered clusters were observed outside of cells through human papillomavirus RNA in situ hybridization detection. Varied numbers of human papillomavirus infective cells were detected by RNAscope assay in different patients even though they were all human papillomavirus high-risk subtype positive discovered by human papillomavirus PCR results. A cell malignancy related long non-coding RNA, TERC, has been detected in seven patient samples. The patient follow-up information was further analyzed with RNAscope results which indicated a combination of RNAscope positive signals of TERC and human papillomavirus high risk signals in more than 10 cells (cytoplasm or nucleus) may connect with cervical lesion fast progression which deserves further studies in the future.C CONCLUSIONS: Taken together, current study has provided an observable clue for gynecologists to evaluate human papillomavirus infection stage and cell malignancy status which may contribute for assessment of cervical disease progression.