CAJ | 학술논문

目的：探讨溴化乙锭（EB）诱导法建立人线粒体DNA（mtDNA）缺失宫颈癌Hela S3细胞，以及再转入线粒体构建融合细胞的可行性，并对转线粒体细胞进行初步分析。方法：采用低剂量（100 ng/mL）EB诱导法建立mtDNA缺失的ρ0 Hela S3细胞，通过普通PCR、荧光定量PCR（qPCR）进行mtDNA缺失鉴定。采用聚乙二醇（PEG）介导细胞融合法，以正常人血小板为mtDNA供体转入ρ0 Hela S3细胞，构建转线粒体细胞（transmitochondrial cybrids），并通过PCR、qPCR和透射电镜观察进行初步分析和鉴定。结果：普通PCR和qPCR结果证实EB诱导法能够成功建立ρ0 Hela S3细胞，同时结合透射电镜证明转线粒体细胞能够恢复线粒体正常结构。结论：采用EB诱导法可成功建立ρ0 Hela S3细胞，且通过细胞融合构建的转线粒体细胞能够恢复mtDNA复制和正常线粒体结构，为研究mtDNA突变与线粒体功能异常相关疾病的关系提供可靠的实验基础。
목적：탐토추화을정（EB）유도법건립인선립체DNA（mtDNA）결실궁경암Hela S3세포，이급재전입선립체구건융합세포적가행성，병대전선립체세포진행초보분석。방법：채용저제량（100 ng/mL）EB유도법건립mtDNA결실적ρ0 Hela S3세포，통과보통PCR、형광정량PCR（qPCR）진행mtDNA결실감정。채용취을이순（PEG）개도세포융합법，이정상인혈소판위mtDNA공체전입ρ0 Hela S3세포，구건전선립체세포（transmitochondrial cybrids），병통과PCR、qPCR화투사전경관찰진행초보분석화감정。결과：보통PCR화qPCR결과증실EB유도법능구성공건립ρ0 Hela S3세포，동시결합투사전경증명전선립체세포능구회복선립체정상결구。결론：채용EB유도법가성공건립ρ0 Hela S3세포，차통과세포융합구건적전선립체세포능구회복mtDNA복제화정상선립체결구，위연구mtDNA돌변여선립체공능이상상관질병적관계제공가고적실험기출。
Objective: To generate mtDNA-depleted Hela S3 cells and the transmitochondrial cybrids and investigate mtDNA replication and mitochondrial structure.Methods: Hela S3 cells were incubated with 100 ng/mL ethidium bromide (EtBr). Polymerase chain reaction (PCR) and quantitative real-time PCR (qPCR) were per-formed in ρ0 Hela S3 cells. Platelet-mediated transmitochondrial cybrids were established on the basis of ρ0Hela S3 cells and identiifed by PCR, qPCR and transmission electron microscopy (TEM).Results: ρ0 Hela S3 cells had a low level of mtDNA replication and distorted cristae of mitochondria, whereas transmotichondrial cybrids had a normal mtDNA replication and regular arranged cristae of mitochondria.Conlusion: The generation of transmitochondrial cybrids provides a viable method for the further research of the effect of mtDNA mutations on mitochondrial respiratory function associated with diseases.