口腔粘膜上皮細胞におけるCEACAM1によるCandida 細胞壁構成成分β-glucanの認識機構
概要
Ꮫ ㄽ ᩥ
ཱྀ⭍⢓⭷ୖ⓶⣽⬊࠾ࡅࡿ CEACAM1 ࡼࡿ
Candida ⣽⬊ቨᵓᡂᡂศ β-glucan ࡢㄆ㆑ᶵᵓ
బஂ㛫 ⨾㞷
ᗈᓥᏛᏛ㝔་ṑ⸆ಖᏛ◊✲⛉
༤ኈㄢ⛬ ་ṑ⸆Ꮫᑓᨷ
ಟᖺᗘ㸦2019 ᖺᗘ㸧
ᣦᑟᩍဨ㸸ຍ⸨ ຌ୍ᩍᤵ
㸦་⣔⛉Ꮫ◊✲⛉ ⏕యᮦᩱᏛ㸧
┠ḟ
Ϩ㸬 ⥴ゝ ͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐ 1
ϩ㸬 ᐇ㦂ᮦᩱ࠾ࡼࡧ᪉ἲ ͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐ 4
1㸬ᇵ㣴⣽⬊
2㸬ཱྀ⭍⢓⭷ୖ⓶⣽⬊ࡢᶞ❧
3㸬Candida albicans ⏕⳦㸪ຍ⇕Ṛ⳦ࡢㄪᩚ
4㸬Candida albicans ࡽࡢ⣽⬊ቨᵓᡂᡂศ β-glucan ྵ᭷ศᏊࡢᢳฟ
5㸬RNA ᢳฟ
6㸬Reverse Transcriptase-Polymerase Chain Reaction ( RT-PCR )
7㸬Real-Time RT-PCR
8㸬⺮ⓑᢳฟ࠾ࡼࡧ࢚࢘ࢫࢱࣥࣈࣟࢵࢸࣥࢢἲ
9㸬Binding Assay
10㸬චỿ㝆
11㸬⣽⬊⺯ගචᰁⰍ
12㸬CEACAM1 ࣀࢵࢡࢲ࢘ࣥᰴࡢస〇
13㸬DCF-DA ἲ
14㸬᰾ෆ⺮ⓑᢳฟ
15㸬୰ᢠయࡢῧຍ
16㸬⤫ィゎᯒ
Ϫ㸬 ⤖ᯝ ͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐ 16
ϫ㸬 ⪃ᐹ ͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐ 22
Ϭ㸬 ⤖ㄽ ͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐ 30
ϭ㸬 ㅰ㎡ ͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐ 32
Ϯ㸬 ཧ⪃ᩥ⊩ ͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐ 33
ϯ㸬 ᅗㄝ᫂ ͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐ 37
ϰ㸬 ⾲࠾ࡼࡧᅗ͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐͐ 42
Ϩ㸬⥴ゝ
ཱྀ⭍ Candida ࡣ㸪Candida ᒓࡼࡿ᪥ぢឤᰁ࡛ 1)㸪ࡑࡢ୰࡛ࡶ Candida
albicans ࡀ᭱ࡶከࡃ᳨ฟࡉࢀ࡚࠸ࡿ 2)㸬Candida albicans ࡣཱྀ⭍⢓⭷ࡢ⾲㠃ࢥ
ࣟࢽ࣮ࢆᙧᡂࡋ࡚╔ࡍࡿ 3)㸬Candida albicans ࡀཱྀ⭍⢓⭷ୖ⓶⣽⬊╔ࡍࡿ
㸪ቑṪࡋ࡞ࡀࡽཱྀ⭍⢓⭷ධࡋ㸪ཱྀ⭍⢓⭷ࢆᦆയࡍࡿ 4)㸬୍᪉㸪ཱྀ⭍⢓⭷ୖ
⓶⣽⬊ࡣ㸪⮬↛ච࠾࠸࡚ Candida albicans ᑐࡍࡿ㜵ᚚࡢ➨୍⥺࡛࠶ࡿ㸬ཱྀ
⭍⢓⭷ୖ⓶⣽⬊ࡣ㸪Candida albicans ⏕⳦᭚㟢ࡉࢀࡿ㸪⅖ᛶࢧࢺ࢝ࣥ
ࡸࢣࣔ࢝ࣥ㸪ᢠ⳦࣌ࣉࢳࢻࢆ⏘⏕ࡋ㸪ึᮇචᛂ⟅ࢆ⾜࠺ 5, 6, 7)㸬ࡲࡓ㸪Candida
albicans ຍ⇕Ṛ⳦᭚㟢ࡋࡓ㝿ࡣ㸪ཱྀ⭍⢓⭷ୖ⓶⣽⬊ࡽ⅖ࡸࢫࢺࣞࢫᛂ⟅
㛵㐃ࡍࡿࡉࡲࡊࡲ࡞㑇ఏᏊࡢⓎ⌧ࡀቑຍࡍࡿ 8)㸬ࡇࢀࡽࡢ⤖ᯝࡣ㸪Candida ⣽
⬊ቨᵓᡂᡂศࢆㄆ㆑ࡍࡿཷᐜయࡀཱྀ⭍⢓⭷ୖ⓶⣽⬊Ꮡᅾࡍࡿࡇࢆ♧၀ࡋ࡚
࠸ࡿ㸬
β-glucan ࡣ㸪Candida albicans ࡸฟⱆ㓝ẕ Saccharomyces cerevisiae ࡢせ࡞⣽
⬊ቨᵓᡂᡂศ࡛࠶ࡿ 9)㸬㏻ᖖ㸪β-glucan ࡣ Candida ⣽⬊ቨࡢෆᒙࢆᵓᡂࡋ㸪ࡑ
ࡢእᒙࡣ࣐ࣥࢼࣥࡀ⨨ࡋ࡚࠸ࡿࡀ 10)㸪in vivo ࠾ࡼࡧ in vitro ࣔࢹࣝ࠾࠸
࡚ Candida ࣂ࢜ࣇ࣒ࣝᙧᡂ β-glucan ࡣ⣽⬊⾲㠃㟢ฟࡋ࡚࠸ࡿࡢ
ሗ࿌ࡀ࠶ࡿ 11)㸬ࡲࡓ㸪β-glucan ࡣ㸪β-glucan ཷᐜయࢆࡋ࡚༢⌫ࡸ࣐ࢡࣟࣇ࣮
1
ࢪ࡞ࡢච⣽⬊ࡢචᛂ⟅ࢆάᛶࡍࡿࡇࡀ▱ࡽࢀ࡚࠸ࡿ
12)
㸬ࡇࢀࡲ࡛ࡢ
◊✲ࡼࡾ㸪β-glucan 㢛⢏ࡀ ROS/p38MAPK/Nrf2 ࢆࡋཱྀ࡚⭍⢓⭷ୖ⓶⣽⬊ࡢ
ࢫࢺࣞࢫᛂ⟅㑇ఏᏊ Heme oxygenase-1 ( HO-1 ) ࡢⓎ⌧ࢆቑຍࡉࡏ㸪Candida
albicans ឤᰁࡼࡗ࡚⏕ࡌࡿ⣽⬊ෆ㓟ࢫࢺࣞࢫᑐࡍࡿᐟ㜵ᚚ㛵㐃ࡍࡿ
ࡇࢆ᫂ࡽࡋࡓ㸬ࡉࡽ㸪ཱྀ⭍⢓⭷ୖ⓶⣽⬊࡛ࡣ Dectin-1 ࡸ TLR2 ࡞ࡢ
β-glucan ཷᐜయࡀⓎ⌧ࡋ࡚࠸ࡿࡇࢆ☜ㄆࡋࡓࡀ㸪ཱྀ⭍⢓⭷ୖ⓶⣽⬊࠾࠸࡚
ࡇࢀࡽࡢཷᐜయࢆࣀࢵࢡࢲ࢘ࣥࡋ࡚ࡶ㸪β-glucan ࡼࡿ HO-1 ࡢⓎ⌧ࡣᙳ㡪
ࢆ࠼࡞ࡗࡓ 8)㸬ࡼࡗ࡚㸪ཱྀ⭍⢓⭷ୖ⓶⣽⬊࠾࠸࡚㸪HO-1 ࡢⓎ⌧ㄏᑟᙳ
㡪ࢆ࠼ࡿ β-glucan 㛵㐃ࡍࡿࢭࣥࢧ࣮㛵ࡋ࡚ࡣ㸪࠸ࡲࡔ࡛᫂࠶ࡿ㸬
Carcinoembryonic antigen-related cell adhesion molecules 1 ( CEACAM1, CD66a,
⫹㐨⢾⺮ⓑ㉁ ) ࡣ CEA ࣇ࣑࣮ࣜࡢࢧࣈࢢ࣮ࣝࣉ࡛࠶ࡾ㸪ᩘ✀㢮ࡢ㑇ఏᏊࣇ
࣑࣮ࣜࡢᏑᅾࡀሗ࿌ࡉࢀ࡚࠸ࡿ 13)㸬CEACAM1 ࡣ㸪චᛂ⟅ࡸ⣽⬊᥋╔㸪ࡀ
ࢇ⣽⬊࠾ࡅࡿቑṪࡸ࣏ࢺ࣮ࢩࢫ㛵㐃ࡋࡓࡉࡲࡊࡲ࡞ࢩࢢࢼࣝఏ㐩㐣⛬
ᙳ㡪ࢆ࠼ࡿ⣽⬊㛫ࢥ࣑ࣗࢽࢣ࣮ࢩࣙࣥ㛵ࡋ࡚࠸ࡿࡇࡀሗ࿌ࡉࢀ࡚࠸ࡿ
14)
㸬ࡲࡓ㸪CEACAM1 ࡣࡉࡲࡊࡲ࡞ᚤ⏕≀ཎయࡢ⣽⬊ཷᐜయࡋ࡚ࡶ▱ࡽࢀ
࡚࠾ࡾ㸪⤖ྜࡍࡿᚤ⏕≀ࡋ࡚᾿⳦( Neisseria meningitidis ), Moraxella catarrhalis,
Haemophilus influenzae ࡞ࡀሗ࿌ࡉࢀ࡚࠸ࡿ㸬㏆ᖺࡢሗ࿌ࡼࡿ㸪Klaile ࡽ
ࡣ㸪⭠⢓⭷ୖ⓶⣽⬊ࡢ CEACAM1 Candida albicans ⏕⳦ࡀ⤖ྜࡍࡿࡇࢆⓎ
2
ぢࡋࡓ 15)㸬CEACAM1 ࡣཱྀ⭍⢓⭷ୖ⓶⣽⬊ࡶⓎ⌧ࡋ࡚࠸ࡿࡇࡀሗ࿌ࡉࢀ࡚
࠸ࡿࡀ㸪ࡑࡢⓎ⌧ࡣ⅖ࢆక࠺⤌⧊࠾࠸࡚ඃ࡛࠶ࡗࡓ
16)
㸬ࡋࡋ࡞ࡀࡽ㸪
ཱྀ⭍⢓⭷ୖ⓶⣽⬊࠾ࡅࡿ CEACAM1 ࡀ㸪Candida ⣽⬊ቨᵓᡂᡂศࢆㄆ㆑ࡋ㸪
චᛂ⟅㛵㐃ࡋ࡚࠸ࡿྰࡣ᫂ࡽ࡛ࡣ࡞࠸㸬
ᮏ◊✲࡛ࡣ㸪ཱྀ⭍⢓⭷ୖ⓶⣽⬊࠾ࡅࡿ β-glucan ࡛ㄏᑟࡉࢀࡿචᛂ⟅
CEACAM1 ࡢ㛵ಀࢆ᫂ࡽࡍࡿࡓࡵ㸪Candida albicans Candida ⣽⬊ቨ
ᵓᡂᡂศ β-glucan ࡼࡗ࡚ㄏᑟࡉࢀࡿ CEACAM1 ࡢⓎ⌧ࢆ᳨ウࡋࡓ㸬ࡉࡽ㸪
β-glucan CEACAM1 ࡢ┦స⏝ࡸ㸪 β-glucan ࡛ㄏᑟࡉࢀࡿ HO-1 ࡢⓎ⌧
ࡸࡑࡢࢩࢢࢼࣝఏ㐩⤒㊰ᑐࡍࡿ CEACAM1 ࡢᙳ㡪ࢆ᳨ウࡋࡓ㸬
3
ϩ㸬ᐇ㦂ᮦᩱ࠾ࡼࡧ᪉ἲ
1㸬ᇵ㣴⣽⬊
ᮏ◊✲࡛ࡣ㸪hTERT 㑇ఏᏊࢆᑟධࡋṚࡉࡏࡓṚࣄࢺཱྀ⭍⢓⭷ୖ⓶
⣽⬊ RT7 ࢆ⏝࠸ࡓ
17,18)
㸬RT7 ࡣ⾲⓶ゅ⣽⬊ῧຍᅉᏊࢭࢵࢺ-Gold ( KGM-
GoldTMSingleQuotsTM ; ࣟࣥࢨࢪࣕࣃࣥᰴᘧ♫㸪ᮾி ) ࢆྵࡴ⾲⓶ゅ⣽⬊
ᇶᮏᇵᆅ-Gold↓⾑ΎKGM-GoldTMBasalMedium ; ௨ୗ KBM ␎グࡍࡿ㸹
ࣟࣥࢨࢪࣕࣃࣥᰴᘧ♫㸪ᮾிࢆ⏝࠸㸪5%CO2 Ꮡᅾୗ㸪37Υຍ‵ୗ࡛ᇵ㣴ࡋ
ࡓ㸬࡚ࡢ⥅௦ᇵ㣴ࡣ㸪࢝ࣝࢳ࣮ࣕࣉ࣮ࣞࢺ ( CELLSTAR® , greiner bio-one ;
Duesseldolf, German ) ✀ࡋࡓ⣽⬊ࢆࢺࣜࣉࢩࣥ/ EDTA ⁐ᾮ㸦ࣟࣥࢨࢪࣕ
ࣃࣥᰴᘧ♫㸪ᮾி㸧࡛ฎ⌮ࡋ⣽⬊ࢆᾋ㐟㸪ᅇࡋ࡚⾜ࡗࡓ㸬
2㸬ཱྀ⭍⢓⭷ୖ⓶⣽⬊ࡢᶞ❧
ᮏ◊✲࡛⏝࠸ࡓཱྀ⭍⢓⭷ୖ⓶⣽⬊ࡣ㸪ᇙఅṑࡢᢤṑ⾡࠾࠸࡚ษ㝖ࡋࡓᗣ
࡞ṑ⫗⤌⧊ࢆ⏝࠸࡚㸪CELL CULTURE FLASKS ( greiner bio-one )࡚ึ௦ᇵ㣴
ࢆ⾜ࡗࡓࡶࡢࢆ⏝ࡋࡓ 19)㸬ࡇࢀࡽࡢ⤌⧊ࡢ᥇ྲྀ㛵ࡋ࡚ࡣ㸪ᗈᓥᏛ⌮ጤ
ဨ࡛ᢎㄆࡉࢀࡓࣉࣟࢺࢥ࣮ࣝᚑ࠸ࣥࣇ࢛࣮࣒ࢻࢥࣥࢭࣥࢺࢆᚓࡓ
(approval no. ...