卷柏抑制人類骨肉瘤細胞其轉移和侵襲之機制探討

骨肉瘤，是原發性惡性骨腫瘤中最常見的且惡性程度最高的一種。其好發於15-19歲青少年，是最常見的兒童骨癌。惡性骨腫瘤常出現於長骨，如股骨遠端、脛骨近端、膝蓋等關節部位。當發生遠處轉移時，最常轉移之部位為肺部。在臨床上常發現一旦腫瘤由良性轉變為惡性腫瘤時，病人容易因癌細胞的轉移導致存活率下降。基質金屬水解蛋白酶(Matrix metalloproteinase; MMP)在腫瘤微環境的生成、腫瘤細胞的形成、轉移及血管新生上扮演一重要角色。卷柏(Selaginella tamariscina)以往在民間用於治療慢性氣管炎、出血，近年來更有研究指出其萃取物具有抗癌功效，但關於卷柏在癌症轉移研究上仍不多，因此我們探討卷柏在癌轉移上所扮演的角色及機制為何。首先我們利用人類骨肉瘤細胞株U2OS處理卷柏萃取物後，發現卷柏萃取物對U2OS並無細胞毒性，且在gelatin zymography assay中發現MMP-9及MMP-2蛋白活性有下降的趨勢。而MMP-9及MMP-2在過去研究中被指出為癌轉移一重要指標，所以我們利用傷口癒合試驗(Wound healing assay)來觀察卷柏萃取物對細胞移動能力的影響，結果抑制U2OS細胞的移動。另外，以Boyden chamber assay實驗中證實了卷柏萃取物明顯的降低癌細胞轉移和侵襲能力。最後，我們利用Western blot方式觀察卷柏萃取物對於訊息傳遞路徑的影響，結果發現卷柏萃取物抑制了PI3K、磷酸化Akt與磷酸化p38的表現，且MMP-9與MMP-2內生性抑制劑TIMP-1及TIMP-2有大量表現的情形。綜合以上結果證實卷柏萃取物經由抑制訊息傳遞路徑中的PI3K、磷酸化Akt與磷酸化p38來達到調控U2OS骨癌細胞株的轉移及侵襲能力。希望卷柏萃取物也許能應用於骨癌轉移及治療上。

關鍵字

卷柏；基質金屬蛋白水解酶骨肉瘤

並列摘要

Osteosarcoma is the most common and highly malignant primary tumor of bone with an early onset age of 15 to 19 years old. This malignance usually occurs at long bone and joints like distal femoral and proximal femur while lungs are the predominant site of metastases from osteosarcoma. As clinical observation indicating a shift from benign tumor to malignant neoplasm, the survival rate was decreased as the result of metastasis. It has been well known that matrix metalloproteinases (MMPs) play a critical role in angiogenesis, tumor formation and progression, and even contribute to microenviroment formation. Several recent researches indicate that Selaginella tamariscina, an oriental medicine used to treat chronic tracheitis and bleeding, had certain pharmacological activities including anti-cancer effect. However, its effect for preventing tumor progression has not been studied. Therefore, this study was designed to study the role of Selaginella tamariscina in against tumor metastasis and the detailed mechanism. First of all, the cytotoxic effect analysis indicated that Selaginella tamariscina extract (STE) at the ranges of 0 to 50μg/mL did not alter U2OS cells viability. Meanwhile, protein levels of MMP-2 and MMP-9 were decreased in a dose-dependent manner as shown by zymography assay analysis. From results of wound healing assay, cell migration and invasion assay with boyden chamber, it was shown that STE significantly reduced the motility of U2OS cells in a dose-dependent manner. Furthermore, the effect of STE on the MAPKs pathway and PI3K/Akt signallings were examined to show that STE reduced the protein levels of PI3K, phosphor-Akt and phosphor-p38 as well as MMP-2 and MMP-9, while that of TIMP-1 and TIMP-2, the inhibitor of MMP-2 and MMP-9, were significantly increased. Based on these findings, it was suggested that STE exerted an inhibitory effect on several essential steps of metastasis, including cell migration and invasion, by regulating the activities of PI3K, phosphor-Akt and phosphor-p38. Therefore, STE may be a powerful candidate in developing preventive agents for cancer metastasis.

並列關鍵字

Selaginella tamariscina ； MMPs ； U2OS

參考文獻

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卷柏抑制人類骨肉瘤細胞其轉移和侵襲之機制探討

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