Prostaglandin A2에 의한 간세포암종 세포의 Caspase-비의존적 Apoptosis 유도
Prostaglandin A2 Induces Caspase-independent Apoptosis in Hepatocellular Carcinoma Cells
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목적: Prostaglandin (PG)A2는 사람 간세포암종 세포의 세포사멸을 유도하는 것으로 보고되었으나, 자세한 기전에 대해서는 아직 밝혀지지 않았다. 본 연구는 PGA2에 의한 사람 간세포암종 세포의 세포사멸 유도시 caspase의 활성화 여부를 알아보고자 하였다. 대상과 방법: 사람 간세포암종 세포주인 Hep 3B와 Hep G2 세포에 PGA2를 처리하고 4',6'-diamino-2-phenylindole (DAPI) 염색에 의한 핵의 모양 변화 및 agarose젤 전기영동에 의한 염색체 DNA의 분절을 관찰하였으며, caspase-3의 기질인 poly ADP-ribose polymerase (PARP)의 분절을 immunoblot 방법을 이용하여 관찰하였고, PGA2에 의한 간세포암종 세포사망에 대한 caspase 저해제의 효과를 분석하였다. 결과: PGA2는 Hep 3B와 Hep G2 세포의 성장을 농도와 시간에 의존적으로 억제하였고, 세포사멸의 특징적인 형태 변화인 핵의 농축과 분절, genomic DNA의 분절을 일으켰다. PGA2에 의한 세포사멸 유도시 PARP는 분절되지 않았으며, caspase의 억제제인 z-VAD-Fmk와 z-DEVD-Fmk는 PGA2에 의한 간세포암종 세포사망을 억제하지 못하였다. 결론: 이상의 결과로 PGA2는 caspase-비의존적 경로를 통하여 사람 간세포암종 세포주인 Hep 3B와 Hep G2 세포의 세포사멸을 일으키는 것으로 사료된다.
Abstract
Background/Aims: Prostaglandin (PG) A2 has been reported to inhibit the growth of hepatocellular carcinoma cells via activation of apoptosis, although the molecular mechanisms involved have not been clarified, yet. To investigate the mechanism of the PGA2-induced apoptosis, we analyzed the activation of caspases during the apoptosis of hepatoma cell lines. Methods: Induction of apoptosis by PGA2 in hepatoma cell lines, Hep 3B and Hep G2, was assessed by DAPI staining of nuclei and agarose gel electrophoresis of genomic DNA. The involvement of caspases was analyzed by immunoblot analysis of poly ADP-ribose polymerase (PARP) and by checking the effect of caspase inhibitors on PGA2-induced apoptosis. Results: PGA2 inhibited the growth of Hep 3B and Hep G2 cells, accompanying nuclear condensation and fragmentation, and genomic DNA laddering, which are the hallmarks of apoptosis. The PARP was not cleaved during the apoptosis of Hep 3B and Hep G2 cells and caspase inhibitors such as z-VAD-Fmk and z-DEVD-Fmk exerted no effect on the PGA2-induced apoptosis. Conclusions: These results suggest that PGA2 induces apoptosis in Hep 3B and Hep G2 cells via caspase-independent pathway.
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