Alternative TitleCell Motility Factors Involved in Invasion and Metastasis of Human Oral Squamous Cell Carcinoma
Periodical title平成10(1998)年度 科学研究費補助金 基盤研究(C) 研究成果報告書概要 = 1998 Fiscal Year Final Research Report Summary
Note (General)金沢大学医学部附属病院
口腔扁平上皮癌における癌細胞の運動能と浸潤様式との関連を明らかにする目的で浸潤様式の異なる3種類の口腔扁平上皮癌細胞株(OSC20,3型;OSC19,4C型;HOC313,4D型)を用いて細胞の運動能を比較検討するとともに,それらの運動能に関与する運動促進因子の発現と機能について検討した.金コロイド法を用いて細胞の運動性を測定したところ,血清存在下での運動能はHOC313細胞が最も高く、次いでOSC19細胞,OSC20細胞の順であったが,無血清培地ではHOC313細胞のみが大きな運動能を示した.この結果からHOC313細胞が運動促進因子を自ら産生していることが推測された.これらの細胞における自己分泌型運動促進因子(autocrine motility factor.AMF)遺伝子の発現を検討した結果,いずれの細胞でもmRNAの存在が認められたがHOC313細胞で特に強く発現していた.またAMFに対する免疫組織化学染色ではHOC313細胞のみにAMF蛋白の存在が認められ,また抗AMF抗体により運動性が抑制されたことから,HOC313細胞が自己運動促進因子としてAMFを産生していることが明らかになった.扁平上皮癌細胞がAMFを産生・分泌しているのを証明したのは本研究が初めてである.AMFレセプターのmRNAの発現はいずれの細胞にも認められたが,レセプター蛋白の存在量はHOC313細胞>OSC20細胞の順に高く、OSC19細胞では認められなかった.AMFと同一アミノ酸配列を有するグルコース6-リン酸イソメラーゼ(glucose 6-phosphate isomerase,G6PI)を作用させると,HOC313細胞とOSC20細胞は濃度依存的に運動性が亢進し,これはAMFレセプター蛋白の発現量と相応していた.分散因子/肝細胞増殖因子(scatter factor/hepatocyte growth factor,SF/HGF)に対しては,HOC313細胞とOSC20細胞で濃度依存的な遊走性の亢進が認められた.フィブロネクチン(fibronectin,Fn)およびビトロネクチン(vitoronectin,Vn)に対する遊走性はHOC313細胞で著しく,OSC20細胞では認めなかった.以上より,口腔扁平上皮癌細胞の運動能と各種運動促進因子に対する感受性は浸潤様式によって異なり,特に4D型の癌細胞は自己分泌型の運動促進因子を産生・分泌して自らの運動能を高め,周囲組織へび漫性に浸潤していくものと考えられた.
The present study was conducted to clarify the mechanism that determines the mode of invasion of human oral squamous cell carcinomas. Three cell lines derived from human oral squamous cell carcinomas with different modes of invasion were assayed for their ability to migrate or invade in vitro. The cell lines employed were HOC3 13, OSO 19 and OSC2O cells, which respectively derive from oral tumors of the highest (grade 4D), high (grade 40) and moderate (grade 3) invasiveness, A phagokinesis assay with gold particles demonstrated that, among the three cell lines, the rate of migration in serum-containing media was ranked in the following order : H0C313 * OSCI9 > OSC2O.In serum free media, H0C313 cells also exhibited a high motility rate, whereas OSCl9 and OSC2O cells migrated to much lesser extents, suggesting that the HOC313 cells produce motility factors that act on themselves in an autocrine manner. Next, the nature of this activity was investigated. Northern blot analysis revealed that the gene coding for autocrine motility factor (AMF) was extremely strongly expressed in HOC313 cells ; two prominent hybridizing bands were detected at 2.0 and 4.0 kb. On the other hand, OSC19 cells expressed the 2.0 kb mRNA species at a low level, and OSC19 cells had barely detectable amounts of AMF mRNA.AMF proteins were clearly marked by immunohistochemical staining with anti-AMF antibodies in H0C313 cells, but not in OSC19 and OSC2O cells ; the immunoreactivity was localized in pseudopodia in a granular pattern. mRNA encoding AMF receptor was detected at a high level in H0C313 and OSC20 cells, and at a low level in OSCl9 cells. In Western blotting, AMF receptor proteins (gp78) were detected at a high level in H0C313 cells, at a low level in OSC2O cells, and not at all in OSCl9 cells. Relative amounts of the AMF and AMF receptor proteins in three cell lines were well correlated with their motility rates. The migration of HOC313 cells in serum-free media was markedly blocked by anti-AMF antibodies. Further, exogenously added glucose 6-phosphate isomerase, which is known to have the same amino acid sequence as AMF, stimulated HOC313 cell motility in a dose-dependent manner. These results indicate that AMF would at least partly account for the high motility of HOC313 cells, and this is the first demonstration that squamous carcinoma cells produce an autocrine factor that induces their own motility. The present study also established the effects of hepatocyte growth factor (HGF)/scatter factor (SF) and cxtracellular matrix components such as fibronectin and vitronectin on the chemotaxis of the oral squamous carcinoma cells. HGF/SF stimulated in a dose dependent manner the H0C313 cell migration through 8 mum pore sized filter to the highest extent. OSC2O cells exhibited a moderate migration toward HGF/SF, while OSCl9 cells did not respond to it. Fibronectin and vitronectin were also found to stimulate the migration of H0C313 cells. The migration of OSC19 cells was
研究課題/領域番号:09672041, 研究期間(年度):1997 – 1998
出典：研究課題「口腔扁平上皮癌の浸潤・転移に関与する運動因子の解析 」課題番号09672041（KAKEN：科学研究費助成事業データベース（国立情報学研究所）） （https://kaken.nii.ac.jp/ja/report/KAKENHI-PROJECT-09672041/096720411998kenkyu_seika_hokoku_gaiyo/）を加工して作成
Related Materialhttps://kaken.nii.ac.jp/search/?qm=00215013
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-09672041/
https://kaken.nii.ac.jp/ja/report/KAKENHI-PROJECT-09672041/096720411998kenkyu_seika_hokoku_gaiyo/
Data Provider (Database)国立情報学研究所 : 学術機関リポジトリデータベース（IRDB）（機関リポジトリ）