Sunitinib promotes apoptosis via p38 MAPK activation and STAT3 downregulation in oral keratinocytes

概要

ㄽ ᩥ ඲ ᩥ せ ⣙

Sunitinib promotes apoptosis via p38 MAPK
activation and STAT3 downregulation
in oral keratinocytes
㸦ศᏊᶆⓗ⸆ࢫࢽࢳࢽࣈࡣ S0$3. άᛶ࡜ 67$7
ᢚไࢆ௓ࡋཱྀ࡚⭍⢓⭷ୖ⓶⣽⬊࡟࢔࣏ࢺ࣮ࢩࢫ
ࢆㄏᑟࡍࡿ㸧

୺ᣦᑟᩍဨ㸸ຍ⸨ ຌ୍ ᩍᤵ
㸦་⣔⛉Ꮫ◊✲⛉ ⏕యᮦᩱᏛ㸧
๪ᣦᑟᩍဨ㸸┦ᕝ ཭┤ ᩍᤵ
㸦་⣔⛉Ꮫ◊✲⛉ ཱྀ⭍እ⛉Ꮫ㸧
๪ᣦᑟᩍဨ㸸ኴ⏣ ⪔ྖ ᩍᤵ
㸦་⣔⛉Ꮫ◊✲⛉ බ⾗ཱྀ⭍ಖ೺Ꮫ㸧

῝⏣ ⩧ᖹ
㸦་ṑ⸆ಖ೺Ꮫ◊✲⛉ ་ṑ⸆Ꮫᑓᨷ㸧

ศᏊᶆⓗ⸆ࢫࢽࢳࢽࣈࡣ㸪⾑⟶ෆ⓶ቑṪᅉᏊཷᐜయ㸦VEGFR ࢱ࢖ࣉ 1 ࠾ࡼࡧ 2㸧
㸪⾑ᑠᯈ⏤᮶
ቑṪᅉᏊཷᐜయ㸦PDGFR-α ࠾ࡼࡧ PDGFR-β㸧࡞࡝ࡢࢳࣟࢩࣥ࢟ࢼ࣮ࢮཷᐜయࡢ㜼ᐖ๣࡛㸪㌿⛣
ᛶ⭈⣽⬊⒴ᝈ⪅ࡢ➨୍㑅ᢥ⸆࡜ࡋ࡚ᗈࡃ౑⏝ࡉࢀ࡚࠸ࡿࠋ୍᪉㸪ࢫࢽࢳࢽࣈࢆᢞ୚ࡉࢀࡓᝈ⪅ࡢ
10㸣࠿ࡽ 50㸣࡟ཱྀෆ⅖㸪ṑ⫗ฟ⾑㸪⢓⭷἞⒵୙඲㸪࿡ぬ㞀ᐖ㸪ཱྀ⭍஝⇱࡞࡝㸪ᵝࠎ࡞ཱྀ⭍࡟㛵㐃
ࡍࡿ᭷ᐖ஦㇟ࢆㄆࡵࡿࡇ࡜ࡀ▱ࡽࢀ࡚࠸ࡿࠋ୰࡛ࡶཱྀෆ⅖ࡣࢫࢽࢳࢽࣈࢆᢞ୚ࡉࢀ࡚࠸ࡿᝈ⪅
࡟㧗㢖ᗘ࡛Ⓨ⑕ࡋ㸪௚ࡢࢳࣟࢩࣥ࢟ࢼ࣮ࢮཷᐜయ㜼ᐖ๣࡜ẚ㍑ࡋ࡚ቑᝏࡢࢢ࣮ࣞࢻࡀ㧗࠸ࡇ࡜
ࡀሗ࿌ࡉࢀ࡚࠸ࡿࠋࡋ࠿ࡋ࡞ࡀࡽࢫࢽࢳࢽࣈࡀཱྀ⭍⢓⭷࡟ཬࡰࡍ⣽⬊㞀ᐖࡢ࣓࢝ࢽࢬ࣒࡟㛵ࡍ
ࡿሗ࿌ࡣ࡞࠸ࠋࡑࡇ࡛ᮏ◊✲ࡣ㸪ཱྀ⭍⢓⭷࡬ࡢࢫࢽࢳࢽࣈࡢᙳ㡪ࢆ᫂ࡽ࠿࡟ࡍࡿࡓࡵ㸪ཱྀ⭍⢓⭷
ୖ⓶⣽⬊࡟࠾ࡅࡿࢫࢽࢳࢽࣈࡢ⣽⬊യᐖࡸ࢔࣏ࢺ࣮ࢩࢫㄏᑟ࡟ᑐࡍࡿᙳ㡪㸪࠾ࡼࡧࡑࢀࡽࡢࢩ
ࢢࢼࣝఏ㐩⤒㊰࡟ࡘ࠸࡚ゎᯒࡋࡓࠋ
 ࡲࡎ㸪୙Ṛ໬ཱྀ⭍⢓⭷ୖ⓶⣽⬊ RT7 ࡜ึ௦ᇵ㣴ཱྀ⭍ୖ⓶⣽⬊࡟ࢫࢽࢳࢽࣈࢆྵࡴྛ✀ࢳࣟࢩ
ࣥ࢟ࢼ࣮ࢮཷᐜయ㜼ᐖ๣ࢆῧຍࡋ㸪⣽⬊ୖΎ࠿ࡽங㓟⬺Ỉ⣲㓝⣲㸦LDH㸧ࡢᨺฟ࡟ࡼࡿ⣽⬊㞀ᐖ
ࢆ ᐃࡋࡓࠋࡲࡓ RT7 ࡟ࢫࢽࢳࢽࣈࢆῧຍࡋ㸪 TUNEL ἲ࡟ࡼࡗ࡚ᰁⰍࡋ㸪⺯ග㢧ᚤ㙾࡜ࣇࣟ
࣮ࢧ࢖ࢺ࣓ࢺ࣮ࣜ࡟࡚᩿∦໬ࡉࢀࡓ DNA ࢆ᳨ฟࡋࡓࠋࢫࢽࢳࢽࣈࡢῧຍ࡟ࡼࡗ࡚ㄏᑟࡉࢀࡿ࢔
࣏ࢺ࣮ࢩࢫ㛵㐃ࢱࣥࣃࢡࡢⓎ⌧ࢆ࢚࢘ࢫࢱࣥࣈࣟࢵࢸ࢕ࣥࢢ ἲ࡟᳨࡚ウࡋࡓࠋFITC ᶆ㆑
AnnexinV/PI ᰁⰍ࡛ྠᐃࡉࢀࡓ࢔࣏ࢺ࣮ࢩࢫ⣽⬊ࡢẚ⋡ࢆࣇ࣮ࣟࢧ࢖ࢺ࣓ࢺ࣮ࣜ࡟ࡼࡗ࡚ゎᯒࡋ
ࡓࠋࡉࡽ࡟ࢫࢽࢳࢽࣈῧຍ࡟ࡼࡿ MAPK Family㸪STAT3 ࡢࣜࣥ㓟໬ࡸ㸪ࢫࢽࢳࢽࣈㄏᑟᛶࡢ⣽
⬊㞀ᐖࡸ࢔࣏ࢺ࣮ࢩࢫ࡟ᑐࡍࡿࡑࢀࡽࢩࢢࢼࣝఏ㐩⤒㊰㜼ᐖ⸆ࡢᙳ㡪ࢆ࢚࢘ࢫࢱࣥࣈࣟࢵࢸ࢕
ࣥࢢࡸ㸪LDH assay, AnnexinV/PI ᰁⰍ࡟᳨࡚ウࡋࡓࠋ
௨ୖࡢᐇ㦂࡟ࡼࡗ࡚㸪௨ୗࡢ⤖ᯝࡀᚓࡽࢀࡓࠋ
1.ࢫࢽࢳࢽࣈࡢῧຍ࡟ࡼࡗ࡚ RT7㸪ึ௦ᇵ㣴ཱྀ⭍ୖ⓶⣽⬊࠿ࡽྛ✀ࢳࣟࢩࣥ࢟ࢼ࣮ࢮཷᐜయ㜼ᐖ
๣ῧຍࡼࡾ㧗࠸ LDH ࡢᨺฟࢆㄆࡵ㸪⣽⬊㞀ᐖࡀ♧ࡉࢀࡓࠋ
2㸬RT7 ࡬ࡢࢫࢽࢳࢽࣈࡢῧຍ࡟ࡼࡗ࡚ DNA ᩿∦໬㸪AnnexinV 㝧ᛶ⣽⬊ࡢቑຍࢆㄆࡵ࢔࣏ࢺ
࣮ࢩࢫࡀㄏᑟࡉࢀࡿࡇ࡜ࡀ♧၀ࡉࢀࡓࠋ
㸬ࢫࢽࢳࢽࣈࡢῧຍ࡟ࡼࡗ࡚࢔࣏ࢺ࣮ࢩࢫ㛵㐃ࢱࣥࣃࢡ࡛࠶ࡿ PARP ࡢⓎ⌧ቑຍ㸪Caspase-3
ࡢάᛶ໬ࢆㄆࡵࡓࠋ୍᪉㸪ᢠ࢔࣏ࢺ࣮ࢩࢫࢱࣥࣃࢡ࡛࠶ࡿ Bcl-2 ࡢⓎ⌧ࡀᢚไࡉࢀࡓࠋࢫࢽࢳࢽ
ࣈ࡟ࡼࡗ࡚࢔࣏ࢺ࣮ࢩࢫෆᅉ⤒㊰㛵㐃ࢱࣥࣃࢡ caspase-9 ࡢάᛶ໬ࡀㄆࡵࡽࢀࡓࡀ㸪እᅉ⤒㊰
㛵㐃ࢱࣥࣃࢡ caspase-8 ࡢάᛶ࡟ኚ໬ࡣㄆࡵࡽࢀ࡞࠿ࡗࡓࠋ
4㸬ࢫࢽࢳࢽࣈࡢῧຍ࡟ࡼࡗ࡚ p38MAPK ࡢࣜࣥ㓟໬ࡀㄏᑟࡉࢀࡓࠋ୍᪉㸪ERK㸪JNK ࡢࣜࣥ
㓟໬ࡣㄆࡵ࡞࠿ࡗࡓࠋp38MAPK 㜼ᐖ๣ SB203580 ࡟ࡼࡿ๓ฎ⌮ࡣ㸪ࢫࢽࢳࢽࣈㄏᑟᛶࡢ
p38MAPK ࡢࣜࣥ㓟໬ࢆᢚไࡋࡓࠋ

5. ࢫࢽࢳࢽࣈࡢῧຍ࡟ࡼࡗ࡚ᐃᖖ≧ែ࡛ㄏᑟࡉࢀࡿ STAT3 ࡢ ࣜࣥ㓟໬ ࡀᢚไࡉࢀࡓࠋ
SB203580 ࡣࢫࢽࢳࢽࣈ࡛ᢚไࡉࢀࡓ STAT3 ࡢࣜࣥ㓟໬ࢆቑຍࡋࡓࠋ
6. SB203580 ࡢ๓ฎ⌮ࡣࢫࢽࢳࢽࣈㄏᑟᛶࡢ LDH ࡢᨺฟࢆᢚไࡋࡓࠋࡉࡽ࡟ SB203580 ࡢ๓
ฎ⌮࡟ࡼࡗ࡚ࢫࢽࢳࢽࣈ࡛ቑຍࡍࡿ࢔࣏ࢺ࣮ࢩࢫ⣽⬊ࡢ๭ྜࡀపୗࡋࡓࠋ
௨ୖࡢ⤖ᯝࡼࡾ㸪ࢫࢽࢳࢽࣈࡣ㸪ཱྀ⭍⢓⭷ୖ⓶⣽⬊࡟࠾ࡅࡿ p38MAPK ࡢࣜࣥ㓟໬ࢆㄏᑟࡋ㸪
STAT3 ࡢάᛶ໬ࢆᢚไࡍࡿࡇ࡜࡟ࡼࡗ࡚㸪⣽⬊㞀ᐖࡸ࢔࣏ࢺ࣮ࢩࢫࢆㄏᑟࡍࡿࡇ࡜ࡀ᫂ࡽ࠿࡟
࡞ࡗࡓࠋp38MAPK 㜼ᐖ๣ࡣࢫࢽࢳࢽࣈ࡟ࡼࡿ⣽⬊㞀ᐖࡸ࢔࣏ࢺ࣮ࢩࢫࡢㄏᑟࢆᢚไࡍࡿࡇ࡜
ࡀ♧ࡉࢀࡓࠋࢫࢽࢳࢽࣈ࡟㛵㐃ࡍࡿཱྀෆ⅖࡞࡝ࡢཱྀ⭍ࡢ᭷ᐖ஦㇟ࡣཱྀ⭍⢓⭷ୖ⓶⣽⬊࡟ᑐࡍࡿ
ࢫࢽࢳࢽࣈࡢ⣽⬊㞀ᐖᛶࡸ࢔࣏ࢺ࣮ࢩࢫㄏᑟࡀ㛵㐃ࡋ࡚࠸ࡿྍ⬟ᛶࡀ⪃࠼ࡽࢀࡓࠋ