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口腔癌細胞のシスプラチン耐性化に伴うNa, K-ATPase及びouabain感受性の変化

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Title: 口腔癌細胞のシスプラチン耐性化に伴うNa, K-ATPase及びouabain感受性の変化
Other Titles: Change of Na,K - ATPase activity and ouabain sensitivity in oral cancer cells accompanied by acquisition of cisplatin resistance
Authors: 平川, 直 Browse this author
Keywords: シスプラチン
P - 糖タンパク質
Na,K - ATPase
Issue Date: 25-Mar-2014
Abstract: cis-diamminedichloro-platinum(II)(シスプラチン、CDDP)による癌の化学療法においては癌細胞の耐性化が障害となるが、その機序には不明な点も多い。そこで、口腔癌細胞株(Sa3、H1、KB)及びそれらのCDDP耐性株(Sa3R、H1R、KBR)と正常歯肉から採取した細胞を用いて、口腔癌細胞のCDDP耐性化に伴うNa,K-ATPaseの変化を中心として検討した。CDDPによる50%細胞生存濃度(IC50)は、H1とH1Rではそれぞれ1.5×10-4及び7.5×10-4M、Sa3とSa3Rでは4.5×10-3及び5.5×10-3M、KBとKBRでは1.0×10-4及び7.0×10-4Mであり、正常歯肉細胞は5.0×10-5Mであった。癌細胞は正常歯肉細胞に比べてCDDP感受性が低く、耐性株はそれぞれの感受性株より低い感受性を示した。Na,K-ATPase活性は、Sa3よりSa3Rが高く、H1よりH1Rがわずかに高値を示したが、KBとKBRで差はなかった。ouabain存在下での各細胞のIC50は、H1とH1Rでは5及び20 nM、Sa3とSa3Rでは80及び180 nM、KBとKBRでは40及び130 nMであり、正常歯肉細胞は5 nMであった。この結果から、CDDP耐性化はouabainに対する耐性化も伴うことが示唆された。各親株と耐性株間のNa,K-ATPase活性阻害に対するouabain濃度依存性は細胞によって異なり、ouabainに対する耐性化はNa,K-ATPaseの変化によるものではないと示唆された。各細胞のP-糖タンパク質とATP7Aの発現を親株と耐性株間で比較したところ、発現量は細胞によって異なった。以上の結果から、CDDP耐性化のメカニズムは細胞の種類によって異なることと、CDDP耐性化はouabain耐性化を伴うことが示唆された。
Purpose:Though resistance of cancer cells is an obstacle in the chemotherapy of cancer by cisplatin (CDDP), the mechanism is not well elucidated.We examined the change of Na,K-ATPase accompanied by acquisition of the CDDP resistance of oral cancer cells.Methods:Oral cancer cell lines (Sa3, H1, KB) and CDDP-resistant lines of them (Sa3R, H1R, KBR) were provided by Faculty of Medicine, Chiba University and cells collected from normal gingiva were used. Na, K-ATPase activity and cell viability in under treatment of ouabain and CDDP for each cell were measured. Assessment of Na, K-ATPase α subunits, ATP7A and P- glycoplotain (P-gp) was done by western blotting.Results:CDDP 50% inhibitory concentrations (IC50) were, 7.5 × 10-4 and 1.5 × 10-4M respectively in H1R and H1, 6.0×10-3 and 4.5×10-3M respectively in Sa3R and Sa3, 7.0×10-4 and 1.0×10-4M respectively in KBR and KB. CDDP sensitivity of cancer cells is lower than the normal gingival cells and induced resistant strains showed a lower sensitivity than original cells. Na, K-ATPase activity was measured. Sa3R is higher than Sa3, and H1R was higher than H1 slightly, but there was no significant difference in the KBR and KB. The results of Na, K-ATPase activity were consistent with protein expression level. Then IC50 of each cell in the presence of ouabain were measured and they were 80 and 60 nM in the H1R and H1, 180 and 80 nM in the Sa3R and Sa3, 130 and 40 nM in the KBR and KB, and normal gingival cells was 5 nM. These results suggest CDDP resistant cells are also resistant to ouabain. The ouabain concentration dependency of Na,K-ATPase inhibition of cells were measured. The 50% inhibitory concentration between resistant and parent lines were different in the cells tested. Therefore, there seemed no relation between the sensitivities of the cells and Na,K-ATPase activities to ouabain. It was suggested that the mechanism of the change of CDDP senseitivity were different in the cells and P-gp, Na,K-ATPase and ATP7A are related to it.
Conffering University: 北海道大学
Degree Report Number: 甲第11271号
Degree Level: 博士
Degree Discipline: 歯学
Examination Committee Members: (主査) 教授 鄭 漢忠, 教授 鈴木 邦明, 教授 進藤 正信
Degree Affiliation: 歯学研究科(口腔医学専攻)
Type: theses (doctoral)
Appears in Collections:学位論文 (Theses) > 博士 (歯学)
課程博士 (Doctorate by way of Advanced Course) > 歯学院(Graduate School of Dental Medicine)

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