Snake venom from Vipera lebetina turanica inhibits tumor in a PC-3 cell Xenograft model and PC-3 cell growth in vitro.

다국어 초록 (Multilingual Abstract)

Objectives : We in here investigated whether snake venom toxin (SVT) from Vipera lebetina turanica inhibits cell growth of human prostate cancer cells (PC-3) Xenograft model and PC-3 cell growth in vitro through induction of apoptosis.
Material and Methods : Mass and weight of he PC-3 human prostate cancer cell that was treated with SVT were evaluated on apoptosis assays, Immunofluorescence staining, confocal immonocytochemistry, electromobility shift assay and tumorigenecity study in athymic nede mice.
Results :Compared with controls, Mass and weight of he PC-3 cell lessened through the above experiments .
Conclusions : These in vitro findings suggest that nano molar range of SVT from Vipera lebetina turanica could inhibit prostate cancer cell growth, and the effect may be related with the NF-κB signal-mediated induction of apoptosis. In addition to the In vitro results, SVT(0.2, 0.4 mg/Kg) suppressed growth of prostate cancer in a xenograft athymic mice model, and reduced volume and weight of the cancer.
Key words :Key words: snake venom toxin, Vipera lebetina turanica, prostate cancer, apoptosis, NF-κB, p53, caspase-3, caspase-9, Bcl-2, Bax, Xenograft, PC-3

국문 초록 (Abstract)

연구목적 : 이 연구는 Vipera lebetina turanica의 蛇毒藥鍼液(Snake venom toxin, SVT)이 in vitro에서 NF-κB의 활성억제와 apoptosis 관련 단백질의 발현 조절을 통하여 세포자멸사(Apoptosis)를 유도하는지 in vivo에서 또한 전립선 암세포주인 PC-3 세포의 성장을 억제하는지 살펴보고자 하였다.

실험방법 : SVT를 처리한 후 PC-3의 성장억제를 관찰하기 위해 WST-1 assay, CCK-8 assay를 시행하였고, Apoptosis evaluation에는 DAPI, TUNEL staining assay를 시행하였으며, Apoptosis regulatory proteins의 변화 관찰에는 western blot analysis를 시행하였고, apoptosis와 연관된 NF-κB의 활성 변화를 관찰하기 위해 EMSA시행하였으며, SVT의 핵내이동을 관찰하기 위해 Immunofluorescence Staining, Confocal immunocytochemistry를 시행하였으며, 전립암세포의 종양형성에는 흉선을 제거한 쥐에 Tumorigenecity study를 시행하였다.

결 과:
PC-3 세포에 SVT를 처리한후, 전립선암세포의 성장, Apoptosis의 유발, Apoptosis관련 단백질의 발현, NF-κB의 활성, SVT의 PC-3세포 핵내 이동여부 및 흉선제거 후 PC-3 세포를 이식한 쥐의 종양형성과정에 미치는 영향을 관찰하여 다음과 같은 결과를 얻었다.
1. PC-3 세포에서 SVT를 처리한 후 세포성장이 억제되고, 세포자멸사가 유도되며, 조절인자인 p53, caspase-3, -9는 증가되었고, Bcl-2는 감소되었다.

2. PC-3 세포에서 SVT를 처리한 후 NF-κB의 활성이 유의하게 감소되었다.

3. DAPI로 염색된 상태에서 SVT가 PC-3 세포의 핵내로 이동되는 것이 관찰되었다.

4. 흉선 제거 후 전립선 암세포주를 이식한 쥐에서 SVT를 피내로 주입한 결과 전립선암의 크기와 무게가 유의하게 감소하였다.

결 론 : 이상의 결과는 SVT가 NF-κB의 활성 억제를 통하여 인간 전립선암세포주인 PC-3의 세포자멸사를 유발함으로써 증식억제 효과가 있음을 입증한 것이며, 이를 재확인한 생체 연구에서의 긍정적인 결과는 향후 SVT의 전립선암의 예방과 치료에 대한 효과적인 치료제 개발에 초석이 될 것으로 기대된다.

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