사람 유두종바이러스 16형의 E6 및 E7 ORF DNA에 의해 형질변환된 자궁경부 상피세포의 특성

Abstract

[한글]

사람 유두종바이러스 16형은 자궁경부암과 밀접한 연관성이 있는 것으로 알려져 있고, 특히 E6 및 E7은 사람 유두종바이러스 16형이 관계한다고 알려진 생식기 암조직 및 여러 세포주에 잘 보존되어 있다. 사람 유두종바이러스 감염이 없는 정상 keratinocyte임을 확

인한 자궁경부 상피세포의 배양 결과 정상 대조 자궁경부 상피세포는 4회 passage 이후에 세포가 노화되어 소멸하였으나 사람 유두종바이러스 16형의 E6E7 DNA를 유핵세포 expression vector인 pZip neo SV(X)에 삽입시켜 제조한 재조합 pZipE6E7 plasmid DNA를 transfection 시킨 자궁경부 상피세포는 10회의 passage 까지 분열 증식하는 수명 연장 현상을 관찰하였다. 정상 대조 군과 사람 유두종바이러스 16형 E6 및 E7 ORF DNA에 의해 형질변환된 세포에 있어서 성장 속도의 차이는 없었으며 형태학적 변화도 관찰되지 않았다.PCR 반응 및 Southern blot hybridization을 통해 E6 및 E7 ORF DNA에 의해 형질변환된 자궁경부 상피세포에서 transfection시킨 E6E7 DNA가 존재함을 확인할 수 있었다. 이상의 실험 결과로 미루어 사람 유두종바이러스의 E6 및 E7 ORF DNA 조각은 사람 정상 자궁경부

상피세포 불멸화에 중요한 역할을 하는 인자로 사료된다.

Chracterization of transformed normal human cervical keratinocyte by E6E7 DNA of

human papillomavirus type 16

Hyun-young Joo

Department of medical Science The Graduate School, Yonsei University

(Directed by Professor Yoon Soo Kim)

Humman papillomavirus(HPV) type-16 is highly associated with cervical dysplasia

and carcinoma and genital tumors

In this study, normal human cervical keratinocytes free from the tumor associated

human papillomaviruses were prepared from normal cervical tissue and transfected

with pZip-E6E7 of HPV recombinant DNA by lipofectin. After G418 treatment for 48

hours, control cells lost immediately their proliferating ability, while

transfected cells continuously primary culture, while control cells lost the

proliferating ability and became senescent after 4th passage. Transfected cells

showed the extended life span(10 passages, 18 weeks) compared to that or

non-transfected control(4 passages,4 weeks),but their growth rate and morphology

did not change during the experimental period. We have demonstrated that

immortalized cells retained the E6E7 DNA fragments of HPV type 16 PCR-Southern blot

analysis. This study suggests that E6E7 DNA fragments of HPV type 16 by

PCR-Southern blot analysis. This study suggests that E6E7 KNA or HPV type 16 may

play an important role in the immortalization of normal human cervical

keratinocytes.

[영문]

Humman papillomavirus(HPV) type-16 is highly associated with cervical dysplasia and carcinoma and genital tumors

In this study, normal human cervical keratinocytes free from the tumor associated human papillomaviruses were prepared from normal cervical tissue and transfected with pZip-E6E7 of HPV recombinant DNA by lipofectin. After G418 treatment for 48

hours, control cells lost immediately their proliferating ability, while transfected cells continuously primary culture, while control cells lost the proliferating ability and became senescent after 4th passage. Transfected cells showed the extended life span(10 passages, 18 weeks) compared to that or

non-transfected control(4 passages,4 weeks),but their growth rate and morphology did not change during the experimental period. We have demonstrated that immortalized cells retained the E6E7 DNA fragments of HPV type 16 PCR-Southern blot analysis. This study suggests that E6E7 DNA fragments of HPV type 16 by PCR-Southern blot analysis. This study suggests that E6E7 KNA or HPV type 16 may play an important role in the immortalization of normal human cervical keratinocytes.