Department of Molecular Pathology, Shinshu University Graduate School of Medicine, Matsumoto, Japan.
PLoS One. 2013;8(1):e54278. doi: 10.1371/journal.pone.0054278. Epub 2013 Jan 17.
N-Acetylgalactosamine 4-sulfate 6-O-sulfotransferase (GalNAc4S-6ST) is the sulfotransferase responsible for biosynthesis of highly sulfated chondroitin sulfate CS-E. Although involvements of CS-E in neuronal cell functions have been extensively analyzed, the role of GalNAc4S-6ST in astrocytic tumor progression remains unknown. Here, we reveal that GalNAc4S-6ST transcripts were detected in astrocytic tumors derived from all 30 patients examined using quantitative reverse transcription-PCR analysis. Patients with high GalNAc4S-6ST mRNA expression had significantly worse outcome compared with patients with low expression, and multivariate survival analysis disclosed that GalNAc4S-6ST is an independent poor prognostic factor for astrocytic tumors. We then tested whether CS-E enhanced haptotaxic migration of glioblastoma U251-MG cells that endogenously express both the CS-E's scaffold tyrosine phosphatase ζ (PTPζ) and GalNAc4S-6ST, in the presence of CS-E's preferred ligands, pleiotrophin (PTN) or midkine (MK), using a modified Boyden chamber method. Haptotaxic stimulation of cell migration by PTN was most robust on control siRNA-transfected U251-MG cells, while that enhancing effect was cancelled following transduction of GalNAc4S-6ST siRNA. Similar results were obtained using MK, suggesting that both PTN and MK enhance migration of U251-MG cells by binding to CS-E. We also found that PTPζ as well as PTN and MK were frequently expressed in astrocytic tumor cells. Thus, our findings indicate that GalNAc4S-6ST mRNA expressed by astrocytic tumor cells is associated with poor patient prognosis likely by enhancing CS-E-mediated tumor cell motility in the presence of PTN and/or MK.
N-乙酰半乳糖胺 4-硫酸 6-O-硫酸转移酶（GalNAc4S-6ST）是负责生物合成高度硫酸化的软骨素硫酸 CS-E 的硫酸转移酶。尽管 CS-E 在神经元细胞功能中的作用已经被广泛分析，但 GalNAc4S-6ST 在星形细胞瘤进展中的作用仍然未知。在这里，我们通过定量逆转录-PCR 分析发现，GalNAc4S-6ST 转录本存在于 30 名患者的星形细胞瘤中。GalNAc4S-6ST mRNA 表达水平高的患者与表达水平低的患者相比，预后明显较差，多变量生存分析显示 GalNAc4S-6ST 是星形细胞瘤的独立不良预后因素。然后，我们使用改良 Boyden 室法测试了 CS-E 是否增强了内源性表达 CS-E 支架酪氨酸磷酸酶 ζ（PTPζ）和 GalNAc4S-6ST 的神经胶质瘤 U251-MG 细胞的趋化性迁移，CS-E 的首选配体是血小板衍生生长因子（PTN）或中期因子（MK）。PTN 对对照 siRNA 转染的 U251-MG 细胞的趋化刺激作用最强烈，而转染 GalNAc4S-6ST siRNA 后，该增强作用被取消。使用 MK 得到了类似的结果，表明 PTN 和 MK 均可通过与 CS-E 结合来增强 U251-MG 细胞的迁移。我们还发现 PTPζ 以及 PTN 和 MK 均在星形细胞瘤细胞中频繁表达。因此，我们的研究结果表明，星形细胞瘤细胞表达的 GalNAc4S-6ST mRNA 可能通过增强 PTN 和/或 MK 存在下的 CS-E 介导的肿瘤细胞迁移与患者预后不良相关。
