[논문]교모세포종 암줄기세포에 대한 진피 소수성 추출물의 항암 활성

김유진; 심예은; 정혜진

doi:10.9721/kjfst.2022.54.1.28

교모세포종 암줄기세포에 대한 진피 소수성 추출물의 항암 활성
Anticancer activity of chloroform extract of Citrus unshiu Markovich peel against glioblastoma stem cells 원문보기

한국식품과학회지 = Korean journal of food science and technology, v.54 no.1, 2022년, pp.28 - 34

김유진 (선문대학교 제약생명공학과) , 심예은 (선문대학교 제약생명공학과) , 정혜진 (선문대학교 제약생명공학과)

초록
AI-Helper

본 연구에서는 진피 소수성 추출물(CECU)의 U87MG 교모세포종 암줄기세포에 대한 항암 활성을 확인하였다. 그 결과, CECU는 25-200 ㎍/mL의 농도 범위에서 U87MG 교모세포종 암줄기세포의 증식, 종양구체 형성과 이동능력을 유의적으로 저해하였다. 특히, CECU는 G0/G1기에서 세포주기 정지와 세포사멸을 유도하여 교모세포종 암줄기세포의 증식을 억제할 수 있었다. 게다가, 교모세포종 암줄기세포에 대한 CECU의 항암 활성은 CD133, Oct4, Nanog, Integrin α6, ALDH1A1과 같은 줄기세포능 조절인자들의 발현과 STAT3 신호전달경로를 저해함으로써 유도된 것임을 확인하였다. 마지막으로, CAM assay를 통해 CECU가 U87MG 교모세포종 암줄기세포의 in vivo 종양 형성을 효과적으로 억제함을 입증하였다. 따라서, 본 연구는 진피 소수성 추출물이 주요 stemness marker들의 발현과 핵심 stemness 조절 신호전달경로를 억제함으로써 U87MG 교모세포종 암줄기세포에 대한 항암 활성을 나타냄을 입증하여, 교모세포종의 예방 및 치료를 위한 천연물 소재로서의 활용 가능성을 새롭게 제시하였다.

Abstract ▼ AI-Helper

Glioblastoma is the most common primary malignant brain tumor and has an extremely poor prognosis. Glioblastoma stem cells (GSCs) contribute to tumor initiation, recurrence, and resistance to therapy, and are thus a key therapeutic target. The peel of Citrus unshiu Markovich has been used in traditional medicine in East Asia to treat various diseases. In this study, we investigated the anticancer activity and molecular mechanism of the chloroform extract of this natural product (CECU) in U87MG GSCs. The results show that CECU inhibited the proliferation, tumorsphere formation, and migration of U87MG GSCs by causing cell cycle arrest at the G0/G1 phase and apoptosis. In addition, CECU downregulated key cancer stemness regulators, including CD133, Oct4, Nanog, integrin α6, ALDH1A1, and STAT3 signaling in U87MG GSCs. Furthermore, CECU significantly suppressed in vivo tumor growth of U87MG GSCs in a chorioallantoic membrane model. Therefore, CECU can be utilized as a natural medicine for the prevention and treatment of glioblastoma.

주제어

표/그림 (6)

그림 Fig. 1. CECU inhibits the proliferation and tumorsphere formation of U87MG GSCs. (A) Effect of CECU on the proliferation of U87MG GSCs. Cells were treated with CECU (12.5-100 μg/mL) and incubated for 7 days. Cell proliferation was measured using the CellTiter-Glo^® luminescent assay. (B) Effect of CECU on the tumorsphere forming ability of U87MG GSCs. Cells were treated with CECU (25-100 μg/mL) and incubated for 7 days. The number and size of tumorspheres in each well were observed under an optical microscope (scale bar: 200 μm). Each value represents the mean±SD of three different experiments. Values are significantly different from the control (***p<0.001).
그림 Fig. 2. Hesperidin and hesperetin inhibit the proliferation and tumorsphere formation of U87MG GSCs. (A) Chemical structures of hesperidin and hesperetin. (B) Effects of hesperidin and hesperetin on the proliferation of U87MG GSCs. Cells were treated with hesperidin and hesperetin (12.5-100 μM) and incubated for 7 days. Cell proliferation was measured using the CellTiter-Glo^® luminescent assay. (C) Effects of hesperidin and heperetin on the tumorsphere forming ability of U87MG GSCs. Cells were treated with hesperidin and hesperetin (25-100 μM) and incubated for 7 days. The number and size of tumorspheres in each well were observed under an optical microscope (scale bar: 200 μm). Each value represents the mean±SD of three different experiments. Values are significantly different from the control (*p<0.05, ***p<0.001).
그림 Fig. 3. CECU inhibits the migration of U87MG GSCs. Cell migration was measured using the wound healing assay. Cells were incubated in the absence or presence of CECU (25, 50 μg/mL) for 24 h. The migrated area was calculated at the indicated time points using ImageJ software (version 1.5: NIH). Dotted white lines indicate the gap at 0 h. Each value represents the mean±SD of three different experiments. Values are significantly different from the control (*p<0.05).
그림 Fig. 4. CECU causes the cell cycle arrest and cellular apoptosis of U87MG GSCs. (A) Effect of CECU on the cell cycle. (B) Effect of CECU on the apoptotic cell death. Cells were treated with CECU (50-200 μg/mL) for 48 h. The cell cycle distribution and apoptotic cell death were evaluated using a Guava^® Muse^® Cell Analyzer with Muse^® Cell Cycle kit and Muse® Annexin V & Dead Cell kit. Each value represents the mean±SD of three different experiments. Values are significantly different from the control (*p<0.05, ***p<0.001).
그림 Fig. 5. CECU downregulates the expression of cancer stemness markers and phosphorylation of STAT3 in U87MG GSCs. (A, B) Cells were treated with CECU (50, 100 μg/mL) for 48 h, and the protein levels were detected by western blot analysis using specific antibodies and were further quantified by densitometry. β-actin levels were used as an internal control. Each value represents the mean±SD of three different experiments. Values are significantly different from the control (*p<0.05, **p<0.01, ***p<0.001).
그림 Fig. 6. CECU suppresses in vivo tumor growth of U87MG GSCs in a CAM model. Fertilized chick eggs were incubated in a humidified incubator at 37°C. At embryonic day 5, U87MG GSCs were mixed with Matrigel in the absence or presence of CECU (100 μg/egg) and implanted onto the CAM surface inside the silicone ring. Seven days later, the CAMs were photographed, formed tumors were retrieved, and tumor weight was calculated. Each value represents the mean±SD of three different experiments. Values are significantly different from the control (***p<0.001).

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