Roomi M Waheed, Ivanov Vadim, Kalinovsky Tatiana, Niedzwiecki Aleksandra, Rath Matthias
Matthias Rath Research Institute, Cancer Division, 1260 Memorex Drive, Santa Clara, CA 95050, USA.
Oncol Rep. 2005 Dec;14(6):1399-404.
Matrix metalloproteinases (MMPs) have been recognized as key players in the degradation of the extracellular matrix (ECM) by migration and proliferation of endothelial cells and their subsequent invasion of the underlying stroma. The prevention of ECM degradation through the inhibition of MMP activity has been shown to be a promising therapeutic approach to block the invasion that occurs during angiogenesis. In previous studies, we demonstrated the anti-tumor effect of a nutrient mixture (NM) containing ascorbic acid, lysine, proline, green tea extract, arginine, N-acetyl cysteine, selenium, copper and manganese on various tumor cell lines in vivo and in vitro. The aim of the present study was to determine whether this mixture has anti-angiogenic effects on human umbilical vein endothelial cells (HUVECs). At near confluence, the HUVEC cell cultures were tested with NM at 0, 10, 50, 100, 500, and 1000 microg/ml in triplicate at each dose for proliferation, migration, MMP expression, and invasion. Cell proliferation was evaluated by MTT assay, invasion potential by Matrigel invasion, MMP expression by gelatinase zymography, and cell migration by a 2 mm-wide scratch in plates. For tube formation, HUVECs were cultured in previously polymerized Matrigel. NM inhibited HUVEC migration, MMP expression and invasion through Matrigel in a dose-dependent manner. Zymography showed a dose-dependent inhibition of MMP-2 expression with virtual total inhibition at a 500 microg/ml concentration. Invasion through Matrigel was totally inhibited at 500 microg/ml NM. NM reduced cell migration by scratch test in a dose-dependent fashion with total inhibition at a 500 microg/ml concentration. NM also inhibited the tube formation of HUVECs, but did not significantly inhibit cell proliferation. These results together with our earlier findings suggest that NM is a relatively non-toxic formulation with anti-angiogenic effects, such as inhibiting vascular tube formation and endothelial cell invasion and migration.
基质金属蛋白酶（MMPs）被认为是内皮细胞迁移和增殖以及随后侵袭下层基质从而导致细胞外基质（ECM）降解的关键因素。通过抑制MMP活性来防止ECM降解已被证明是一种有前景的治疗方法，可阻断血管生成过程中的侵袭。在先前的研究中，我们证明了一种含有抗坏血酸、赖氨酸、脯氨酸、绿茶提取物、精氨酸、N - 乙酰半胱氨酸、硒、铜和锰的营养混合物（NM）在体内和体外对各种肿瘤细胞系具有抗肿瘤作用。本研究的目的是确定该混合物对人脐静脉内皮细胞（HUVECs）是否具有抗血管生成作用。接近汇合时，以0、10、50、100、500和1000微克/毫升的浓度对HUVEC细胞培养物进行NM测试，每个剂量一式三份，以检测增殖、迁移、MMP表达和侵袭情况。通过MTT法评估细胞增殖，通过基质胶侵袭实验评估侵袭潜力，通过明胶酶谱法评估MMP表达，通过在平板上划2毫米宽的划痕评估细胞迁移。对于血管生成，将HUVECs培养在预先聚合的基质胶中。NM以剂量依赖性方式抑制HUVEC迁移、MMP表达和通过基质胶的侵袭。酶谱分析显示对MMP - 2表达有剂量依赖性抑制，在500微克/毫升浓度时几乎完全抑制。在500微克/毫升NM时，通过基质胶的侵袭被完全抑制。通过划痕试验，NM以剂量依赖性方式减少细胞迁移，在500微克/毫升浓度时完全抑制。NM还抑制HUVECs的血管生成，但对细胞增殖没有显著抑制作用。这些结果与我们早期的发现共同表明，NM是一种相对无毒的制剂，具有抗血管生成作用，如抑制血管管形成以及内皮细胞侵袭和迁移。
