Serum Tenascin-X Strongly Binds to Vascular Endothelial Growth Factor
Serum Tenascin-X Strongly Binds to Vascular Endothelial Growth Factor
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DOI：
10.1248/bpb.32.1004
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发表时间：
2009-06-01
影响因子：
2
通讯作者：
Matsumoto, Ken-ichi
中科院分区：
文献类型：
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作者：
Ishitsuka, Taichi;Ikuta, Tomoki;Matsumoto, Ken-ichi
间质细胞外基质tenascin-X(ITNX)大小约为450 kDa，广泛存在于多种组织中。此前，我们在小鼠中鉴定了大小为200 kDa的血清形式的TnX(STnX)。在本研究中，为了研究sTnX的特性和功能，构建了编码重组小鼠sTnX的质粒。作为对照，我们还构建了编码小鼠450 kDa iTNX和编码250 kDa iTNX的质粒，其中缺失了450 kDa iTNX中200 kDa的sTNX区域。在稳定表达每个重组TNX的细胞中，释放到条件培养液中的200 kDa的sTNX的量是释放到条件培养液中的250 kDa的iTNX和450 kDa的iTNX的7倍多。我们以前报道过iTNX的剪接异构体(340-kDa iTNX)与血管内皮生长因子B(VEGF-B)和血管内皮生长因子A(VEGF-A)结合。因此，用免疫共沉淀法检测了VEGF-A和VEGF-B与200-kDa sTNX的结合能力，并与250-kDa iTNX和450-kDa iTNX的结合能力进行了比较。结果表明，与其他iTNX蛋白的结合能力相比，sTNX与血管内皮生长因子-A和血管内皮生长因子-B的结合能力较强。根据5-乙炔基-2‘-脱氧尿苷(EDU)掺入实验结果，我们发现纯化的重组200-kDa sTNX单独或与血管内皮生长因子-A或碱性成纤维细胞生长因子联合使用对增殖的血管内皮细胞(UV Female 2细胞)的DNA合成有微弱的促进作用。这些结果表明，sTNX对内皮细胞具有较弱的增殖活性。
Interstitial extracellular matrix tenascin-X (iTNX) with about 450 kDa is prominently present in various tissues. Previously, we identified the serum form of TNX (sTNX) with 200 kDa in the mouse. In the present study, in order to investigate distinctive features and functions of sTNX, a plasmid encoding the recombinant mouse sTNX was constructed. As a control, we also constructed a plasmid encoding mouse 450-kDa iTNX and a plasmid encoding 250-kDa iTNX, which lacks the region of 200-kDa sTNX from 450-kDa iTNX. In cells stably expressing each recombinant TNX, a more than 7-fold larger amount of 200-kDa sTNX was released into conditioned medium than the amounts of 250-kDa iTNX and 450-kDa iTNX released into the medium. We previously reported that a splice isoform of iTNX (340-kDa iTNX) binds to vascular endothelial growth factor B (VEGF-B) as well as to VEGF-A. Therefore, the ability of VEGF-A and VEGF-B to bind to 200-kDa sTNX was examined by a co-immunoprecipitation assay in comparison with the binding abilities to 250-kDa iTNX and 450-kDa iTNX. It was found that sTNX strongly bound to VEGF-A and VEGF-B, compared with the binding abilities of other iTNX proteins. Based on the results of assays of incorporation of 5-ethynyl-2'-deoxyuridine (EdU), we found that purified recombinant 200-kDa sTNX both alone and in combination with VEGF-A or basic fibroblast growth factor (bFGF) can weakly promote DNA synthesis in proliferating vascular endothelial cells (UV female 2 cells). These results suggest that sTNX possesses weak activity for proliferation of endothelial cells.