中文摘要：

磷酸酶及张力蛋白的同源基因（PTEN）是一种抑癌基因,可以调控细胞的增殖,与癌症的发生和发展息息相关。本研究采用MTT法和流式细胞术分别检测了重组荞麦胰蛋白酶抑制剂（r BTI）对人肝癌细胞株Hep G2细胞的增殖以及周期的影响。免疫荧光及Western印迹法检测了PTEN和p-PTEN的亚细胞定位及蛋白表达的变化。采用q RT-PCR及Western印迹法检测了周期相关蛋白的表达。旨在探究PTEN和p-PTEN在r BTI抑制Hep G2细胞增殖和周期阻滞中的作用。结果表明,r BTI能显著抑制Hep G2细胞增殖,将细胞周期阻滞在G0/G1期,并呈时间和剂量依赖性;r BTI作用于Hep G2后,可显著上调PTEN和p-PTEN的表达。同时发现,p-PTEN主要分布于细胞核中,能与核仁发生共定位;周期相关蛋白检测表明,细胞内p53、p21转录水平和蛋白水平均增加。综上所述,r BTI通过上调PTEN的表达,使得细胞周期阻滞于G0/G1期,进而抑制Hep G2细胞的增殖。

英文摘要：

Phosphatase and tensin homologue（ PTEN）,acts as a tumor suppressor gene in mammalian cells,can regulate cell proliferation and is related with the occurrence and development of cancer. This study investigated the roles of PTEN and p-PTEN on proliferation and cell cycle in Hep G2 cells induced by recombinant buckwheat trypsin inhibitor（ r BTI）. The effects of r BTI on the proliferation and cell cycle of Hep G2 cells were measured by MTT assays and flow cytometry. The subcellular localization and the expression of PTEN and p-PTEN were determined by immunofluorescence and Western blot. And the changes of protein involved in cell cycle were detected by q RT-PCR and Western blotting. The results showed that r BTI could significantly inhibit the proliferation of Hep G2 cells and cell cycle was arrested at G0/ G1 phase in a dose- and time-dependent manner. Meanwhile, r BTI significantly increased the expression of PTEN in the cytoplasm and p-PTEN in the nucleus. Further study found that p-PTEN was co-localized with nucleolus in the Hep G2 cells. The assessment of cycle-related protein indicated that the transcription and expression of p53 and p21 were increased after treatment with r BTI. Together all,r BTI inhibits Hep G2 cells proliferation by up-regulating the expression of PTEN to arrest cell cycle in G0/ G1 phase.