Koga H, Sakisaka S, Harada M, Takagi T, Hanada S, Taniguchi E, Kawaguchi T, Sasatomi K, Kimura R, Hashimoto O, Ueno T, Yano H, Kojiro M, Sata M
Second Department of Medicine, Kurume University School of Medicine, Kurume, Japan.
Hepatology. 2001 May;33(5):1087-97. doi: 10.1053/jhep.2001.24024.
Peroxisome proliferator-activated receptor gamma (PPARgamma) regulates cell growth and differentiation. Recent evidence has suggested that PPARgamma ligands had anti-tumor effects through inhibiting cell growth and inducing cell differentiation in several types of malignant neoplasm. In the present study, we investigated: 1) the expression of PPARgamma in both human hepatoma cell lines and 5 resected human hepatocellular carcinoma (HCC) tissues; 2) the growth-inhibitory effect of troglitazone, a PPARgamma ligand, on those hepatoma cells; and 3) the molecular mechanisms of troglitazone-induced cell-cycle arrest. Five hepatoma cell lines, HLF, HuH-7, HAK-1A, HAK-1B, and HAK-5, were used. The mRNA expression levels of PPARgamma, p21(WAF1/Cip1), and p27(Kip1) were determined by real-time quantitative reverse transcription-polymerase chain reaction. The expression of cell cycle-regulating proteins, such as p21, p27, p18(INK4c), cyclin E, and pRb, was examined using Western blotting. PPARgamma was constitutively expressed in all the cell lines and the HCC tissues used in this study. A cytostatic effect of troglitazone was found in those cell lines, and this inhibition of cell growth was dosage-dependent. G0/G1 arrest was apparently demonstrated in flow cytometric analysis in HLF, HAK-1A, HAK-1B, and HAK-5, all of which showed an increased expression of p21 protein. However, HuH-7, lacking p21 protein expression, did not demonstrate clear arrest in the cell-cycle analysis. HLF, which was deficient in the protein product of the retinoblastoma tumor-suppressor gene (pRb), responded most profoundly to troglitazone, showing an increased expression in not only p21, but also in p27 and in p18. These findings suggested that p21, p27, and p18 might be involved in troglitazone-induced cell-cycle arrest in human hepatoma cells.
过氧化物酶体增殖物激活受体γ（PPARγ）调节细胞生长和分化。最近有证据表明，PPARγ配体通过抑制几种恶性肿瘤类型的细胞生长和诱导细胞分化而具有抗肿瘤作用。在本研究中，我们调查了：1）PPARγ在人肝癌细胞系和5例手术切除的人肝细胞癌（HCC）组织中的表达；2）PPARγ配体曲格列酮对这些肝癌细胞的生长抑制作用；3）曲格列酮诱导细胞周期停滞的分子机制。使用了5种肝癌细胞系，即HLF、HuH-7、HAK-1A、HAK-1B和HAK-5。通过实时定量逆转录-聚合酶链反应测定PPARγ、p21（WAF1/Cip1）和p27（Kip1）的mRNA表达水平。使用蛋白质印迹法检测细胞周期调节蛋白如p21、p27、p18（INK4c）、细胞周期蛋白E和pRb的表达。PPARγ在本研究中使用的所有细胞系和HCC组织中组成性表达。在这些细胞系中发现了曲格列酮的细胞生长抑制作用，并且这种对细胞生长的抑制是剂量依赖性的。在HLF、HAK-1A、HAK-1B和HAK-5的流式细胞术分析中明显显示出G0/G1期停滞，所有这些细胞系均显示p21蛋白表达增加。然而，缺乏p21蛋白表达的HuH-7在细胞周期分析中未显示出明显的停滞。视网膜母细胞瘤肿瘤抑制基因（pRb）的蛋白产物缺陷的HLF对曲格列酮反应最强烈，不仅p21表达增加，p27和p18表达也增加。这些发现表明，p21、p27和p18可能参与曲格列酮诱导的人肝癌细胞的细胞周期停滞。
