양막성분으로 배양한 각막섬유모세포에서의 Differential Gene Expression

목적 : 배양된 인간 각막 섬유모세포에서, 염증반응, 세포고사, 세포분화 및 성장에 관련된 유전자 중심으로 양막성분의 세포보호효과 및 유전자 발현의 변화를 알아보았다.<BR> 대상과 방법 : 양막을 액화질소속에서 분쇄하고 4℃에서 균질화 한 후, 원심분리 하여 상층액을 얻었다. 각막 섬유모세포를 tumor necrosis factor-α(TNF-α)와 양막성분을 첨가한 경우와 TNF-α만을 첨가한 경우로 나누어 배양하여 각 군에서의 세포의 형태와 생존율을 확인하였고, cDNA Array 방법을 이용하여 유전자 발현 변화를 측정하였다.<BR> 결과 : 양막성분과 TNF-α를 함께 처치한 군이 TNF-α만을 처치한 군에 비하여 세포생존율이 높고, 세포의 형태변화가 적었다. cDNA array상 양막성분을 처리한 군에서 interleukin(IL)-1β, IL-1 receptor type 1의 발현과 caspase-1,-3,-10은 발현이 감소되었으며, nuclear factor-κB(NF-κB), IκB kinase α, integrin-α1과 -α10, mitogen-activated protein(MAP) kinase 및 matrix metalloproteinases(MMPs)는 발현이 증가되었다.<BR> 결론 : 배양된 각막섬유모세포에 TNF-α자극시, 양막성분은 염증유발유전자와 세포고사 유전자의 발현을 억제하고, 세포의 분화 및 성장을 촉진 시키는 유전자의 발현을 증가시킴으로써 세포를 보호한다고 생각된다.〈한안지 42(12):1760-1770, 2001〉

Purpose : To know whether amniotic membrane (AM) extract could protect cell from toxic stimuli by changing gene expression such as inflammatory and apoptosis-related genes, cultured corneal fibroblasts were used.<BR> Methods : After AM was pulverized in the liquid nitrogen and homogenized under the 4(C, it was centrifuged and its supernatant was obtained. Human corneal fibroblasts (HCFs) were divided into 2 groups - one group treated with AM extracts and TNF-α, and the other with TNF-αonly. Cell morphology and viability were assessed and differential gene expression through cDNA array was performed.<BR> Results : Viability of HCF cultured in AM extracts and TNF-αwas better than that of cells in TNF-αonly. Cells in TNF-αonly were morphologically changed and dead. cDNA array showed the decrease in IL-1ß, IL-1 receptor type 1 and caspases-1,-3,and -10 expression in cells treated with AM extracts. MMP-1,-2,-3 &-9 were almost the same or slightly increased and NF-κB, IκB kinase α, integrin-α1 and -α10, and MAP kinase were increased in cells with AM extracts.<BR> Conclusions : When cultured corneal fibroblasts were insulted by TNF-α, AM extracts seemed to protect the cells, by down-regulating various inflammation - and cell death- related genes, and by increasing the expression of genes enhancing cell proliferation and differentiation. J Korean Ophthalmol Soc 42(12):1760-1770, 2001