[논문]매실(Prunus mume) 메탄올 분획물의 처리에 따른 인체 전립선암세포 LNCaP의 apoptosis 유도 효과

김휘곤; 김정호; 허지안; 원영선; 서권일

doi:10.5352/jls.2021.31.3.321

매실(Prunus mume) 메탄올 분획물의 처리에 따른 인체 전립선암세포 LNCaP의 apoptosis 유도 효과
Induction of Apoptosis by Treatment of Human Prostate Cancer LNCaP Cells with Methanol Fractions from Prunus mume 원문보기

생명과학회지 = Journal of life science, v.31 no.3, 2021년, pp.321 - 329

김휘곤 (동아대학교 응용생명과학과) , 김정호 (경북대학교 식품공학부) , 허지안 (동아대학교 식품생명공학과) , 원영선 (동아대학교 식품생명공학과) , 서권일 (동아대학교 식품생명공학과)

초록
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본 연구에서는 매실 메탄올 추출물(maesil methanol fraction, MMF)을 제조하여 인체 전립선암세포 LNCaP, RC-58T 및 PC-3에 대한 증식억제 효과를 확인하였다. 인체 전립선암세포인 PC-3 및 RC-58T와 비교해보았을 때, LNCaP은 MMF의 처리에 따른 증식억제 효과에 가장 민감했다. LNCaP의 형태학적 관찰과 apoptotic body 형성을 관찰해보았으며, MMF의 처리로 인한 형태의 변화, 핵 손상 및 응축을 확인했다. MMF의 처리로 인한 인체 전립선암세포 LNCaP에서 성장억제 효과가 내인성 apoptosis 경로와 관련 있는지 확인한 결과, pro-apoptotic 단백질인 Bax, caspase-3, caspase-9, PARP의 발현이 증가하였고, anti-apoptotic 단백질인 Bcl-2의 발현이 감소하는 것을 확인했다. MMF와 AIF inhibitor인 N-phenylmalemide (N-PM)의 병용처리군에 비해 MMF 단독처리군의 증식억제 효과가 유의적으로 나타났으며 AIF 및 Endo G의 발현 증가를 통해 외인성 apoptosis 경로에 영향을 미치는 것을 확인했다. 또한 PI3K inhibitor인 LY294002와 MMF의 병용처리군에 비해 MMF 단독처리군의 증식억제 효과가 유의적으로 나타났으며 PI3K, p-Akt, p-mTOR의 발현 감소를 통해 PI3K/Akt/mTOR 신호경로에 영향을 미치는 것을 확인했다. 결론적으로 인체 전립선암세포 LNCaP에서 MMF의 증식억제 효과는 천연물 유래 기능성 식품의 소재로써의 가능성을 보여준다.

Abstract ▼ AI-Helper

This study examined the growth inhibitory effect of the methanol fraction of maesil (Prunus mume) extract (MMF) on LNCaP, PC-3, and RC-58T human prostate cancer cell lines. Among these cell lines, LNCaP was the most sensitive to the inhibitory effects of MMF. Observation of the morphology and apoptotic body formation in the LNCaP cells revealed morphological changes, nuclear damage, and condensation in response to MMF treatment. The suppressive effect of MMF was related to the intrinsic apoptosis pathway, as indicated by increased expression of the pro-apoptotic proteins Bax, capase-3, capase-9, and PARP and decreased expression of the anti-apoptotic protein Bcl-2. Combined treatment with MMF and the AIF inhibitor N-phenylmalemide (N-PM) indicated that MMF treatment alone had a significant growth suppression effect. The involvement of the extrinsic apoptosis pathway was also confirmed by increased expression of AIF and Endo G. The growth suppression effect of MMF was also significant when compared to the effects of a combination of the PI3K inhibitor LY294002 and MMF. The reduced expression of PI3K, p-Akt, and p-mTOR confirmed the involvement of the PI3K/Akt/ mTOR signaling pathway in regulating the anti-proliferative properties of MMF. In conclusion, the growth suppression effect of MMF in the LNCaP human prostate cancer cell line shows the possibility of using this natural product in functional foods.

주제어

표/그림 (6)

$Fig . 1. Cell growth inhibitory effect on human prostate cancer cell. (A) RC-58T, LNCaP and PC-3 treated with MMF for 24 hr by SRB assay. (B) LNCaP treated with MMF for 24 hr by SRB assay. Data values are expressed as mean ± SD of triplicate determinations. Significant differences were compared with control at *p<0.05, **p<0.01 and ***p<0.001 by student’s t-test. MMF: maesil methanol fraction.$ 그림 Fig . 1. Cell growth inhibitory effect on human prostate cancer cell. (A) RC-58T, LNCaP and PC-3 treated with MMF for 24 hr by SRB assay. (B) LNCaP treated with MMF for 24 hr by SRB assay. Data values are expressed as mean ± SD of triplicate determinations. Significant differences were compared with control at *p<0.05, **p<0.01 and ***p<0.001 by student’s t-test. MMF: maesil methanol fraction.
$Fig. 2. Effect of MMF on morphological change of human prostate cancer LNCaP cells. Cell morphology was visualized by optical microscopy (×200). MMF: maesil methanol fraction.$ 그림 Fig. 2. Effect of MMF on morphological change of human prostate cancer LNCaP cells. Cell morphology was visualized by optical microscopy (×200). MMF: maesil methanol fraction.
$Fig. 3. Effect of MMF on nuclear fragmentation in human prostate cancer LNCaP cells. Cells were stained with 10 μg/ml of Hoechst 33,258 and examined by fluorescene. MMF: maesil methanol fraction.$ 그림 Fig. 3. Effect of MMF on nuclear fragmentation in human prostate cancer LNCaP cells. Cells were stained with 10 μg/ml of Hoechst 33,258 and examined by fluorescene. MMF: maesil methanol fraction.
$Fig. 4. Effect of MMF on intrinsic apoptosis pathway in human prostate cancer LNCaP cells. Cells were treated with MMF for 24 hr. Bax, Bcl-2, cleaved PARP, caspase-3 and caspase-9 was assessed by Western blot analysis. Cell lysates were subjected to 12% SDS-PAGE. MMF: maesil methanol fraction.$ 그림 Fig. 4. Effect of MMF on intrinsic apoptosis pathway in human prostate cancer LNCaP cells. Cells were treated with MMF for 24 hr. Bax, Bcl-2, cleaved PARP, caspase-3 and caspase-9 was assessed by Western blot analysis. Cell lysates were subjected to 12% SDS-PAGE. MMF: maesil methanol fraction.
$Fig. 5. Effect of MMF on extrinsic apoptosis pathway in human prostate cancer LNCaP cells. (A) The influence of N-PM (AIF inhibitor) was measured by SRB assay. (B) AIF and Endo G was assessed by Western blot analysis. Data values are expressed as mean ± SD of triplicate determinations. Significant differences were compared with control at *p<0.05, **p<0.01 and ***p<0.001 by student's t-test. Cells were treated with MMF for 24 hr. Cell lysates were subjected to 12% SDS-PAGE. MMF: maesil methanol fraction.$ 그림 Fig. 5. Effect of MMF on extrinsic apoptosis pathway in human prostate cancer LNCaP cells. (A) The influence of N-PM (AIF inhibitor) was measured by SRB assay. (B) AIF and Endo G was assessed by Western blot analysis. Data values are expressed as mean ± SD of triplicate determinations. Significant differences were compared with control at *p<0.05, **p<0.01 and ***p<0.001 by student's t-test. Cells were treated with MMF for 24 hr. Cell lysates were subjected to 12% SDS-PAGE. MMF: maesil methanol fraction.
$Fig. 6. Effect of MMF on PI3K/Akt/mTOR pathway in human prostate cancer LNCaP cells. (A) The influence of LY294002 (PI3K inhibitor) was determined by SRB assay. (B) PI3K, Akt, p-Akt, mTOR and p-mTOR was assessed by Western blot analysis. Data values are expressed as mean ± SD of triplicate determinations. Significant differences were compared with control at *p<0.05, **p<0.01 and ***p<0.001 by student's t-test. Cells were treated with MMF for 24 hr. Cell lysates were subjected to 12% SDS-PAGE. MMF: maesil methanol fraction.$ 그림 Fig. 6. Effect of MMF on PI3K/Akt/mTOR pathway in human prostate cancer LNCaP cells. (A) The influence of LY294002 (PI3K inhibitor) was determined by SRB assay. (B) PI3K, Akt, p-Akt, mTOR and p-mTOR was assessed by Western blot analysis. Data values are expressed as mean ± SD of triplicate determinations. Significant differences were compared with control at *p<0.05, **p<0.01 and ***p<0.001 by student's t-test. Cells were treated with MMF for 24 hr. Cell lysates were subjected to 12% SDS-PAGE. MMF: maesil methanol fraction.

AI 본문요약
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문제 정의

이에 본 연구에서는 MMF의 처리에 따른 인체 전립선암세포 성장 억제 효능 및 apoptosis 유도 효과를 확인하였으며이를 기능성 소재로의 이용가능성을 제시하는 기초자료로 활용하고자 하였다.

데이터처리

각 군 간의 통계적 유의성 검증은 stu- dent’s t-test로 판정하였다(*p<0.05, **p<0.01, ***p<0.001).
본 실험에서 이용된 통계분석은 one-way ANOVA의 방법으로 시료군 간의 유의적인 변화를 분석하였으며, 평균 값과 표준편차로 나타내었다. 각 군 간의 통계적 유의성 검증은 stu- dent’s t-test로 판정하였다(*p<0.

성능/효과

MMF의 처리는 인체 전립선암세포 LNCaP의 내인성 및 외인성 apoptosis 경로의 활성화를 통해 증식을 억제하였을 뿐만 아니라 PI3K/Akt/mTOR 신호경로의 조절을 통해 인체 전립선암세포의 성장을 억제하였다. 결론적으로 이번 연구를 통해 MMF의 천연물 유래 전립선암세포 증식 억제 효능을 통해 매실이 갖는 기능성 소재로써의 활용가능성을 제시하였다.

후속연구

MMF의 처리는 인체 전립선암세포 LNCaP의 내인성 및 외인성 apoptosis 경로의 활성화를 통해 증식을 억제하였을 뿐만 아니라 PI3K/Akt/mTOR 신호경로의 조절을 통해 인체 전립선암세포의 성장을 억제하였다. 결론적으로 이번 연구를 통해 MMF의 천연물 유래 전립선암세포 증식 억제 효능을 통해 매실이 갖는 기능성 소재로써의 활용가능성을 제시하였다.

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저자의 다른 논문 :

표제어: PCR

동의어: Packet Collision Rate

용어 설명 출처 목록 (6)

용어 설명: PCR은 세균 특이성이 있는 primer를 이용하여 적은 수의 세균이 있을지라도 쉽게 검출할 수 있는 유용한 방법이며, 이를 이용하여 구강 내 치면세균막이나 타액에서 직접 세균을 검출할 수 있게 되었다[8].

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