생체 외 도입에 따른 단백질의 세포 내 발현 양상 (Intracellular Expression of Proteins After Transduction Into Human Cells in vitro)

서지정보

· 발행기관 : 대한구강악안면병리학회
· 수록지 정보 : 대한구강악안면병리학회지 / 36권 / 6호 / 331 ~ 340페이지
· 저자명 : 임현주, 최종선, 홍승희

초록

Internalization and expression of extracellular molecules into cells and tissues is known very important process to biological processes and therapy of various diseases. In this study, we analyzed expression pattern of extracellular molecule after transduction into various human cells. To investigate cellular expression of internalized molecule, we used adenovirus containing green fluorescence protein. After infection of adenovirus into various human cells, the efficiency of intracellular gene expression was assessed with determining GFP expressing cells by fluorescence microscopy or FACS. After one day of adenovirus infection into HepG2 and A549, we observed that GFP expression was low at 10moi but expression levels were increased at 100moi in both cells. But, adenovirus infection into HCT116 showed low expression of GFP at concentrations from 1moi to 100moi. After 2 day infection with adenovirus, GFP expression level at 10moi and 100moi was highly increased in HepG2 and A549 compared with 1 day infection. Especially, GFP expression was significantly increased in HCT116after 2 days infection. However, GFP expressing SKOV3 cells by adenovirus infection were not found in all the experimental conditions tested. For quantitative analysis of GFP expression of cells by adenovirus infection, we carried out FACS analysis. As a result, GFP was expressed at very low levels at 1moi in all cells used in this experiment. GFP expression slightly increased after increasing moi to 10in HepG2, HCT116, and A549 cells. By 100moi infection of adenovirus, GFP expression was elevated to 10 fold higher than 10moi in HepG2 and A549 and about 4 fold elevation was observed in HCT116. A549 showed 20 fold higher expression of GFP than SKOV3.
We also found that GFP expression by adenovirus infection was the highest in HepG2 cells. Protein expression was enhanced by increasing concentrations or time of adenovirus infection. In these results, GFP expression efficiency of adenoviral gene transduction reveals the highest in HepG2 and lowest in SKOV3 among the cells tested. Taken together, we could confirm that intracellular protein expression efficiency by transduction of extracellular gene was different in various human cells. Our study suggests that the cell types and cellular properties should be carefully examined to enhance expression efficiency of extracelluar molecules in biological research and disease therapy.

영어초록

Internalization and expression of extracellular molecules into cells and tissues is known very important process to biological processes and therapy of various diseases. In this study, we analyzed expression pattern of extracellular molecule after transduction into various human cells. To investigate cellular expression of internalized molecule, we used adenovirus containing green fluorescence protein. After infection of adenovirus into various human cells, the efficiency of intracellular gene expression was assessed with determining GFP expressing cells by fluorescence microscopy or FACS. After one day of adenovirus infection into HepG2 and A549, we observed that GFP expression was low at 10moi but expression levels were increased at 100moi in both cells. But, adenovirus infection into HCT116 showed low expression of GFP at concentrations from 1moi to 100moi. After 2 day infection with adenovirus, GFP expression level at 10moi and 100moi was highly increased in HepG2 and A549 compared with 1 day infection. Especially, GFP expression was significantly increased in HCT116after 2 days infection. However, GFP expressing SKOV3 cells by adenovirus infection were not found in all the experimental conditions tested. For quantitative analysis of GFP expression of cells by adenovirus infection, we carried out FACS analysis. As a result, GFP was expressed at very low levels at 1moi in all cells used in this experiment. GFP expression slightly increased after increasing moi to 10in HepG2, HCT116, and A549 cells. By 100moi infection of adenovirus, GFP expression was elevated to 10 fold higher than 10moi in HepG2 and A549 and about 4 fold elevation was observed in HCT116. A549 showed 20 fold higher expression of GFP than SKOV3.
We also found that GFP expression by adenovirus infection was the highest in HepG2 cells. Protein expression was enhanced by increasing concentrations or time of adenovirus infection. In these results, GFP expression efficiency of adenoviral gene transduction reveals the highest in HepG2 and lowest in SKOV3 among the cells tested. Taken together, we could confirm that intracellular protein expression efficiency by transduction of extracellular gene was different in various human cells. Our study suggests that the cell types and cellular properties should be carefully examined to enhance expression efficiency of extracelluar molecules in biological research and disease therapy.

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