Detection of human papillomavirus type 16 early-gene transcription by reverse transcription-PCR is associated with abnormal cervical cytology

Abstract

Human papillomavirus type 16 (HPV-16) is associated with abnormal Papanicolou smears, indicative of cervical intraepithelial neoplasia. HPV-16 is the most common genital HPV and is found in up to 40% of young women with normal cervical cytology. In order to investigate whether transcriptionally active HPV-16 infection is associated with abnormal cervical smears, a reverse transcription-nested PCR assay with primers from the E5 open reading frame was developed to detect all HPV-16 early-region mRNA (E-mRNA) transcripts. It was used to study HPV-16-infected women with normal and abnormal cervical cytologies to obtain evidence of active infection. Among HPV-16 DNA-positive women, HPV-16 E-mRNA was detected in 15 of 37 (40.5%) women with abnormal cervical cytology but in only 4 of 35 (11.4%) women with normal cytology (P ‫؍‬ 0.007). Thus, HPV-16 E-mRNA transcription is associated with abnormal cervical smears and may have value as a prognostic marker of progressive disease.

Human papillomavirus 16/18 E5 promotes cervical cancer cell proliferation, migration and invasion in vitro and accelerates tumor growth in vivo

“…Individual colonies of G418-selected cells were isolated and expanded in DMEM/FBS. Cell lines were examined for E5 mRNA expression using an adaptation reverse transcriptase-polymerase chain reactions (RT-PCR) described by Biswas et al (11). The cells were tested for the presence of the GFP-E5 fusion protein (no commercial E5 antibody) by western blot analyses as described below.…”

Analyses of variant human papillomavirus type-16 E5 proteins for their ability to induce mitogenesis of murine fibroblasts

“…Cell lines were tested for E5 mRNA expression using an adaption of the method described by Biswas et al . [ 5 ]. Briefly reverse transcriptase (RT) reactions were primed using the HPV-16 E5 downstream primer ( 4110 TACAGGATCCTTATGTA ATTAAAAAGCGTGCAT 4078 ) with Moloney murine leukemia virus reverse transcriptase then samples were subjected to a nested PCR using internal primers located within the E5 ORF [ 5 ].…”

HPV16/18 E5, a promising candidate for cervical cancer vaccines, affects SCPs, cell proliferation and cell cycle, and forms a potential network with E6 and E7

“…Total-RNA was reverse-transcribed by the RT-PCR System (Life Technologies, USA) (12). The primer sets used are shown in Table II (generated by Takara Biotechnology Co., Ltd., Japan).…”