Abe T, Mori T, Kohno K, Seiki M, Hayakawa T, Welgus H G, Hori S, Kuwano M
Department of Biochemistry, Kyushu University School of Medicine, Japan.
Clin Exp Metastasis. 1994 Jul;12(4):296-304. doi: 10.1007/BF01753836.
Metalloproteinases, inhibitors of metalloproteinases, plasminogen activators, inhibitors of plasminogen activators and cathepsins are thought to be involved in invasion by tumor cells. Glioblastoma multiforme is highly malignant and extremely refractory to therapy. One reason is because of its highly invasive nature within the nervous system. However, it remains unclear how invasion/dissemination of glioblastoma multiforme proceeds. In this study, we attempted to determine which proteinases were responsible for the invasion activity of human glioma cell lines in vitro. Nine human glioma cell lines (NHG1, NHG2, IN157, IN301, IN500, U251, U343, T98G and CCF-STTG1) derived from patients with glioma were grown in culture and used. We compared the invasion activity of glioma cell lines in a Matrigel invasion assay system, and formulated the activity as invasion index (%). Among the nine cell lines, IN157, IN500 and U343 showed less than 10% invasion activity (low group); NHGI, IN301 and CCF-STTG1 showed 10-25% activity (intermediate group); NHG2, U251 and T98G showed more than 30% activity (high group). Addition of an inhibitor of metalloproteinases, TIMP-1, to the assay system was found to significantly inhibit invasion activity of T98G cells (P < 0.01). Northern blot analysis demonstrated expression of urokinase-type plasminogen activator (uPA), tissue-type PA (tPA) and PA inhibitor-1 (PAI-1) in some of the above cell lines. Cellular levels of PAs and their inhibitor mRNA, however, appeared not to be correlated with invasion activity in most glioma cell lines except for CCF-STTG1. Expression of 72 kDa type IV collagenase (MMP-2) was much lower in IN157, IN500 and U343 than other cell lines, whereas expression of TIMP-1 was much higher in IN500 than in other cell lines. Zymographic activity was found to be comparable to MMP-2 mRNA levels in all cell lines except for CCF-STTG1. Type IV collagenolytic activity was also comparable to invasion activity in nine cell lines. These observations suggest the role of type IV collagenase and its inhibitors in determining capacity for invasion by human gliomas. However, a comprehensive analysis both in vitro and in vivo is required to confirm the role for this enzyme in glioma cell invasiveness.
金属蛋白酶、金属蛋白酶抑制剂、纤溶酶原激活剂、纤溶酶原激活剂抑制剂和组织蛋白酶被认为与肿瘤细胞的侵袭有关。多形性胶质母细胞瘤恶性程度高，治疗极其困难。原因之一是其在神经系统内具有高度侵袭性。然而，多形性胶质母细胞瘤的侵袭/扩散过程仍不清楚。在本研究中，我们试图确定哪些蛋白酶负责体外培养的人胶质瘤细胞系的侵袭活性。使用了来自胶质瘤患者的9个人胶质瘤细胞系（NHG1、NHG2、IN157、IN301、IN500、U251、U343、T98G和CCF-STTG1）进行培养。我们在基质胶侵袭试验系统中比较了胶质瘤细胞系的侵袭活性，并将该活性表示为侵袭指数（%）。在这9个细胞系中，IN157、IN500和U343的侵袭活性低于10%（低侵袭组）；NHG1、IN301和CCF-STTG1的侵袭活性为10%-25%（中等侵袭组）；NHG2、U251和T98G的侵袭活性高于30%（高侵袭组）。在试验系统中加入金属蛋白酶抑制剂TIMP-1后，发现可显著抑制T9G细胞的侵袭活性（P<0.01）。Northern印迹分析表明，上述部分细胞系中存在尿激酶型纤溶酶原激活剂（uPA）、组织型PA（tPA）和PA抑制剂-1（PAI-1）的表达。然而，除CCF-STTG1外，大多数胶质瘤细胞系中PA及其抑制剂mRNA的细胞水平似乎与侵袭活性无关。IN157、IN500和U343中72kDa IV型胶原酶（MMP-2）的表达远低于其他细胞系，而IN500中TIMP-1的表达远高于其他细胞系。除CCF-STTG1外，所有细胞系中的酶谱活性均与MMP-2 mRNA水平相当。IV型胶原酶活性在9个细胞系中也与侵袭活性相当。这些观察结果提示IV型胶原酶及其抑制剂在决定人胶质瘤侵袭能力方面的作用。然而，需要在体外和体内进行全面分析，以证实该酶在胶质瘤细胞侵袭性中的作用。
