Guan Ming, Pang Chi-Pui, Yam Hin-Fai, Cheung Kin-Fai, Liu Wei-Wei, Lu Yuan
Center of Laboratory Medicine, Huashan Hospital, Fudan University, Shanghai 200040, PR China.
Cancer Gene Ther. 2004 May;11(5):325-32. doi: 10.1038/sj.cgt.7700675.
Pigment epithelium-derived factor (PEDF) is a potent inhibitor of angiogenesis and an inducer of neural differentiation. We previously reported the loss of PEDF expression in glioma progression. In this study, we investigated whether PEDF overexpression could suppress glioma growth and invasion. Glioma cell line U251 was stably transfected with a full-length human PEDF expression vector. The expression and release of various cytokines and angiogenic factors into the medium were analyzed by real-time reverse transcription-polymerase chain reaction, enzyme-linked immunosorbent assay, and gelatin zymography. Apoptosis was checked by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling. Growth inhibition was evaluated by using the in vitro Matrigel invasion. Tumorigenicity was examined in vivo by subcutaneous xenotransplantation into severe combined immunodeficient mice. In U251 cells overexpressing PEDF, thrombospondin-1 protein was upregulated (5.3-fold more), but the production of vascular endothelial growth factor (VEGF) (1.8-fold less) and basic fibroblast growth factor (2.5-fold less) was lower than in cells transfected with the vector only. PEDF also downregulated the production of matrix metalloproteinase-9. Conditioned medium collected from the PEDF-transfected U251 cells showed a significant reduction of VEGF expression. In vitro invasiveness was reduced by approximately 40%. PEDF expression prevented the growth of transfected cells and caused a significant increase in the percentage of cells undergoing apoptosis (50.4% in PEDF-transfected cells). Furthermore, the size of xenotransplants was significantly smaller. In conclusion, PEDF overexpression decreased malignancy, and this might be attributed to the promotion of apoptosis and the regulation of expression of angiogenic effectors. Thus, treatment with PEDF may be useful in patients with malignant gliomas. However, the mechanism of apoptosis induction needs to be investigated.
色素上皮衍生因子（PEDF）是一种强大的血管生成抑制剂和神经分化诱导剂。我们之前报道过在胶质瘤进展过程中PEDF表达缺失。在本研究中，我们调查了PEDF过表达是否能抑制胶质瘤的生长和侵袭。用全长人PEDF表达载体稳定转染胶质瘤细胞系U251。通过实时逆转录 - 聚合酶链反应、酶联免疫吸附测定和明胶酶谱法分析各种细胞因子和血管生成因子在培养基中的表达和释放。通过末端脱氧核苷酸转移酶介导的dUTP缺口末端标记法检测细胞凋亡。使用体外基质胶侵袭实验评估生长抑制情况。通过皮下异种移植到严重联合免疫缺陷小鼠体内来检测体内致瘤性。在过表达PEDF的U251细胞中，血小板反应蛋白 -1蛋白上调（多5.3倍），但血管内皮生长因子（VEGF）的产生（少1.8倍）和碱性成纤维细胞生长因子（少2.5倍）低于仅用载体转染的细胞。PEDF还下调基质金属蛋白酶 -9的产生。从转染PEDF的U251细胞收集的条件培养基显示VEGF表达显著降低。体外侵袭性降低了约40%。PEDF表达抑制了转染细胞的生长，并导致凋亡细胞百分比显著增加（转染PEDF的细胞中为50.4%）。此外，异种移植瘤的大小明显更小。总之，PEDF过表达降低了恶性程度，这可能归因于促进细胞凋亡和调节血管生成效应因子的表达。因此，PEDF治疗可能对恶性胶质瘤患者有用。然而，细胞凋亡诱导的机制需要进一步研究。
