探討抗菌胜肽m2163與m2386誘導人類腸癌細胞凋亡之路徑 與死亡受器表現

Antimicrobial peptides m2163 and m2386 may interact with the death receptor on the cell surface of SW480 cells and trigger the apoptotic process

廖怡舜(Yi-Shun Liao)

指導教授 ： 林志侯

國立清華大學/生命科學院/分子醫學研究所/碩士(2013年)

摘要

近年來，抗菌胜肽的研究除了抑菌之外，抗癌的潛力越來越受到重視與投入。本實驗室從Lactobacillus casei ATCC 334菌株的基因序列篩選出兩個具有抗菌活性的胜肽m2163與m2386。在過去的研究發現，兩種抗菌胜肽除了可以抑制李斯特菌(Listeria monocytogenes)的生長。並且可以誘發腸道癌細胞SW480進行細胞凋亡，但所造成的細胞凋亡路徑沒有很清楚。本次實驗利用Real time PCR技術測定目標基因的mRNA表現量，預測可能的凋亡路徑，並且用西方墨點法測定各種與細胞凋亡相關的蛋白表現量。實驗結果顯示，以2163與m2386處理人類腸癌細胞SW480後，誘發細胞的FasR或TNFR1等死亡受器活化，接者死亡受器下游的caspase 8也被活化，證實細胞會以死亡受體介導途徑造成細胞凋亡。此外，caspase 8會造成Bid活化成tBid，tBid誘使Bax作用在粒線體上，造成粒線體釋出cytochrome c與Smac等凋亡相關蛋白，證實細胞凋亡也會走向粒線體介導途徑。綜合以上的結果，推論m2163與m2386可能會直接或間接啟動相關受器，或接在細胞膜上造成細胞壓力，引起細胞凋亡的發生。

關鍵字

抗菌胜肽 ； 腸癌細胞 ； 細胞凋亡路徑 ； 死亡受器

並列摘要

In recent years, some of the antimicrobial peptides (AMPs), which are toxic to bacteria, exhibit a strong cytotoxic activity in cancer cells, but not to normal mammalian cells. This intriguing feature has obvious anticancer potential. In the past, our laboratory screened to have active AMPs(m2163 and m2386) from Lactobacillus casei ATCC 334. The AMPs were confirmed to have anticancer activity in human colorectal cancer cell lines, SW480, and the AMPs would induce apoptosis of SW480.But the apoptotic pathways are not very clear. To find out the apoptotic pathways, we used real time PCR technology to test the relative mRNA expression of target gene. And used western blot to test the expression of target protein. The results show that AMPs could induce the expression of death receptor (Fas, TNFR1 and TRAILR1) of SW480 afterward, active the mitochondria release apoptotsis- associated proteins (cytochrome c and Smac). It is confirmed that AMP treatment cause SW480 apoptotic pathways included death receptor-mediated pathway and mitochondria-mediated pathway. These results suggest that the m2163 and m2386 interact with apoptotsis- associated receptor or affect on cell membrane surfaces.

並列關鍵字

antimicrobial peptides ； colorectal cancer ； apoptotic pathways ； death receptor

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