無血清培養系を用いた口腔癌由来細胞株の樹立ならびに機能解析

概要

Ꮫ ఩ ㄽ ᩥ
↓⾑Ύᇵ㣴⣔ࢆ⏝࠸ࡓཱྀ⭍⒴⏤᮶⣽⬊ᰴࡢ
ᶞ❧࡞ࡽࡧ࡟ᶵ⬟ゎᯒ

Ꮫ఩⏦ㄳ⪅ బ⸨ ᡂ⣖
ᗈᓥ኱Ꮫ኱Ꮫ㝔་ṑ⸆ಖ೺Ꮫ◊✲⛉་ṑ⸆Ꮫᑓᨷ
2019 ᖺᗘ

୺ᣦᑟᩍဨ㸸ᒸᮏ ဴ἞ ᩍᤵ
ศᏊཱྀ⭍་Ꮫ࣭㢡㢦㠃እ⛉Ꮫ

ㅰ㎡

ᮏ◊✲࡟㝿ࡋࠊᚚ᠓⠜࡞ࡿᚚᣦᑟ࡞ࡽࡧ࡟ᚚᰯ㜀ࢆ㈷ࡾࡲࡋࡓᗈᓥ኱Ꮫ኱Ꮫ㝔་⣔⛉Ꮫ◊

✲⛉ ศᏊཱྀ⭍་Ꮫ࣭㢡㢦㠃እ⛉Ꮫ ᒸᮏဴ἞ᩍᤵ࡟῝ཌ࡞ࡿㅰពࢆ⾲ࡋࡲࡍࠋࡲࡓࠊᮏᩥస

ᡂୖࠊ⤊ጞᚚ᠓ษ࡞ࡿᚚᣦᑟࠊᚚᰯ㜀ࢆ㈷ࡾࡲࡋࡓᮏᏛ ⏕యᮦᩱᏛㅮᗙ ຍ⸨ຌ୍ᩍᤵࠊ㢡

㢦㠃ゎ๗Ꮫ ᑎᒣ㝯ྖᩍᤵ࡟῝ㅰ⮴ࡋࡲࡍࠋᮏㄽᩥసᡂ࡟㝿ࡋᚚຓゝࠊᚚᰯ㜀ࢆ㈷ࡾࡲࡋࡓࠊ

ᮏᏛṑ⛉ᨺᑕ⥺Ꮫ ᰠᮏ┤ஓᩍᤵࠊᮏᏛබ⾗ཱྀ⭍ಖ೺Ꮫ ኴ⏣⪔ྖᩍᤵࠊᮏᏛศᏊཱྀ⭍་Ꮫ࣭

㢡㢦㠃እ⛉Ꮫ ⹡㇂ⱱ᫛෸ᩍᤵ࡟ឤㅰࡢពࢆ⾲ࡋࡲࡍࠋᑦࠊᮏ◊✲ࢆ㐍ࡵࡿ࡟㝿ࡋࠊ⤊ጞᚚ᠓

ษ࡞ࡿᚚᣦᑟ࡜ᚚ㓄៖ࢆ㈷ࡾࡲࡋࡓᮏᏛ་⣔⛉Ꮫ◊✲⛉ ศᏊཱྀ⭍་Ꮫ࣭㢡㢦㠃እ⛉Ꮫ ℈⏣

඘Ꮚ༤ኈ࡟ᚰ࠿ࡽឤㅰࡢពࢆ⾲ࡋࡲࡍࠋࡉࡽ࡟ࠊከ኱࡞ࡿᚚᨭ᥼ࠊᚚ༠ຊࢆ㡬ࡁࡲࡋࡓᮏᏛ་

ṑ⸆ಖ೺Ꮫ◊✲⛉ ศᏊཱྀ⭍་Ꮫ࣭㢡㢦㠃እ⛉Ꮫᩍ⫋ဨྛ఩࡟ឤㅰࡋࡲࡍࠋ᭱ᚋ࡟ࠊᖖ࡟⮬ศ

ࢆᨭ࠼࡚ࡃࢀࡓᐙ᪘ཬࡧぶ᪘࡟ᚰࡼࡾឤㅰ࠸ࡓࡋࡲࡍࠋ

 










┠ḟ

␎グ   ࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭1

➨ 1 ❶  ⥴ゝ   ࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭4
➨ 2 ❶  ཱྀ⭍⒴⏤᮶⣽⬊ᰴࡢᶞ❧࡞ࡽࡧ࡟ᶵ⬟ゎᯒ
➨ 1 ⠇ ᐇ㦂ᮦᩱཬࡧ᪉ἲ ࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭7
(1) ᇵ㣴ᮦᩱཬࡧᇵ㣴᪉ἲ
(2) ⣽⬊ᰴࡢᶵ⬟ゎᯒ
1) ձ ⣽⬊ቑṪ⬟ࡢ᳨ウ
ղ ONEC-TS ࡢ⣽⬊᥋╔⬟ࡢ᳨ウ
2) ᰁⰍయゎᯒ
3) ࢾ࣮ࢻ࣐࢘ࢫ⫼㒊⓶ୗ࡛ࡢ㐀⭘⒆⬟ࡢ᳨ウ
4) ࣐࢖ࢥࣉࣛࢬ࣐ឤᰁヨ㦂
5) Short tandem repeat (STR) ゎᯒ
6) ⺯ගච␿ᰁⰍἲࢆ⏝࠸ࡓྛ⣽⬊࡟࠾ࡅࡿ EGFR ࡢⓎ⌧᳨ウ
7) EGF ཬࡧᢠ EGFR ᢠయࡢ⣽⬊ቑṪ࡟ཬࡰࡍᙳ㡪
8) ONEC ࡟࠾ࡅࡿୖ⓶࣭⚄⤒⣔࣐࣮࣮࢝ཬࡧ⚄⤒ෆศἪ࣐࣮࣮࢝ࡢⓎ⌧

ゎᯒ

ձ ⺯ගච␿ᰁⰍἲ࡟ࡼࡿୖ⓶⣔ཬࡧ⚄⤒⣔࣐࣮࣮࢝ࡢⓎ⌧᳨ウ

ղ ච␿⤌⧊ᰁⰍἲ࡟ࡼࡿ⚄⤒ෆศἪ࣐࣮࣮࢝ࡢⓎ⌧᳨ウ
ճ Western blot ἲ࡟ࡼࡿ⚄⤒ෆศἪ࣐࣮࣮࢝ࡢⓎ⌧᳨ウ
         9) Transcriptome ゎᯒ

       

10) ྛ⣽⬊ᰴࡢᢠ⒴๣࡟ᑐࡍࡿឤཷᛶヨ㦂
11) PonatinibࠊEverolimusࠊSunitinib ࡢྛ⣽⬊ᰴࡢቑṪཬࡧࢩࢢࢼ

ࣝఏ㐩࡟ཬࡰࡍᙳ㡪
12) qRT-PCR ἲཬࡧ Western blot ἲࢆ⏝࠸ࡓྛ⣽⬊ᰴ࡟࠾ࡅࡿ FGFR

ࡢⓎ⌧᳨ウ
➨ 2 ⠇ ⤖ᯝ  ࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭21
1) ᶞ❧⣽⬊ᰴࡢ఩┦ᕪ㢧ᚤ㙾ീ
2) ձ ྛ⣽⬊ᰴࡢ⣽⬊ቑṪ⬟ࡢ᳨ウ⤖ᯝ
ղ ONEC-TS ࡢ⣽⬊᥋╔⬟ࡢ᳨ウ⤖ᯝ
3) ᰁⰍయゎᯒ⤖ᯝ
4) ࢾ࣮ࢻ࣐࢘ࢫ⫼㒊⓶ୗ࡛ࡢ㐀⭘⒆⬟ࡢ᳨ウ⤖ᯝ
5) ࣐࢖ࢥࣉࣛࢬ࣐ឤᰁヨ㦂⤖ᯝ
6)

STR ゎᯒ⤖ᯝ

7) ⺯ගච␿ᰁⰍἲࢆ⏝࠸ࡓྛ⣽⬊࡟࠾ࡅࡿ EGFR ࡢⓎ⌧᳨ウ⤖ᯝ

8) EGF ཬࡧᢠ EGFR ᢠయࡢ⣽⬊ቑṪ࡟ཬࡰࡍᙳ㡪ࡢ᳨ウ⤖ᯝ
9) ONEC ࡟࠾ࡅࡿୖ⓶࣭⚄⤒⣔࣐࣮࣮࢝ཬࡧ⚄⤒ෆศἪ࣐࣮࣮࢝ࡢⓎ⌧

ゎᯒ⤖ᯝ
ձ ⺯ගච␿ᰁⰍἲ࡟ࡼࡿ ONEC-TS ࡟࠾ࡅࡿୖ⓶⣔ཬࡧ⚄⤒⣔࣐࣮

࣮࢝ࡢⓎ⌧᳨ウ⤖ᯝ
ղ ච␿⤌⧊ᰁⰍἲ࡟ࡼࡿ ONEC-TS ࡟࠾ࡅࡿ⚄⤒ෆศἪ࣐࣮࣮࢝ࡢ

Ⓨ⌧᳨ウ⤖ᯝ
ճ Western blot ἲ࡟ࡼࡿ ONEC-TS ࡟࠾ࡅࡿ⚄⤒ෆศἪ࣐࣮࣮࢝ࡢ

Ⓨ⌧᳨ウ⤖ᯝ
10) Transcriptome ゎᯒ⤖ᯝ
11) ONEC ࡢᢠ⒴๣࡟ᑐࡍࡿឤཷᛶヨ㦂⤖ᯝ
12) PonatinibࠊEverolimusࠊSunitinib ࡢྛ⣽⬊ᰴࡢቑṪཬࡧࢩࢢࢼࣝఏ

㐩࡟ཬࡰࡍᙳ㡪ࡢ᳨ウ⤖ᯝ
13) qRT-PCR ἲཬࡧ Western blot ἲࢆ⏝࠸ࡓྛ⣽⬊ᰴ࡟࠾ࡅࡿ FGFR ࡢ

Ⓨ⌧᳨ウ
➨ 3 ❶  ⪃ᐹ  ࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭28
➨ 4 ❶  ⥲ᣓ  ࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭38

ཧ⪃ᩥ⊩

ᅗ⾲

࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭ 40

࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭࣭50

␎グ

ABP

Alcian blue and PAS stain

BSA

Bovine Serum Albumin

CD56

CD56 molecule

CDDP

cis-diamminedichloro-platinum(II)

cDNA

Complementary deoxyribonucleic acid

CT

Computed tomography

DAB    

Diaminobenzidine

DAPI     4’, 6-diamidino- 2-phenylindole
DMEM

Dulbecco’s modified Eagle’s medium

DT

Doubling time

EDTA

Ethylene Diamine Tetra Acetic acid

EGF

Epidermal growth factor

EGFR

Epidermal growth factor receptor

FGFR     Fibroblast growth factor receptor
H-E

Hematoxylin and eosin stain

HRP

Horseradish peroxidase

kDa

Kilo Dalton

1

pKRT

Pan keratin

mRNA

Messenger RNA

NET

Neuroendocrine tumor

NGFR

Nerve growth factor receptor

OASCC

Oral adenosquamous cell carcinoma

ONEC

Oral neuroendocrine carcinoma

OSCC

Oral squamous cell carcinoma

PBS

Phosphate-buffered saline

PCR

Polymerase chain reaction

PET-CT

Positron emission tomography

PVDF

Polyvinylidene difluoride

qRT-PCR

quantitative Real time protein chain reaction

RPMI

Roswell park memorial institute media

SCC

Squamous cell carcinoma

SDS

Sodium dodecyl sulfate

STR

Short tandem repeat

TBS

Tris-buffered saline

Tris

Tris(hydroxyl methyl) amino methane

2

TritonX-100

Octylphenol ethoxylate

Tween20

Polysorbate 20

VP-16

Etoposid

5-FU

5-Fluorouracil

3

➨ 1 ❶ ⥴ゝ
Gey࡟ࡼࡾࣄࢺᏊᐑ㢕⒴࠿ࡽୡ⏺ึࡢ⒴⣽⬊ᰴ࡛࠶ࡿHeLa⣽⬊ࡀᶞ❧ࡉࢀ࡚௨᮶ (Gey, et
al., 1952)ࠊከࡃࡢ⒴⣽⬊ᰴࡀᶞ❧ࡉࢀ࡚ࡁࡓ (Eagle, 1955; Rangan, 1972)ࠋཱྀ⭍㡿ᇦ࡟࠾࠸࡚ࠊ
Mooreࡽ࡟ࡼࡿႃ㢌⒴⏤᮶H.Ep3⣽⬊ᰴࡢᶞ❧ (Moore, et al., 1955)ࠊᇼ㉺ࡽ࡟ࡼࡿṑ⫗⒴⏤᮶
Ca9-22⣽⬊ᰴ (ᇼ㉺, 1974)ࠊᐑෆࡽࡢHo-1-u1⣽⬊ᰴ࠾ࡼࡧHo-1-n1⣽⬊ᰴ (ᐑෆ et al., 1985;
Miyauchi, et al., 1988) ࢆࡣࡌࡵ࡜ࡋ࡚ࠊከࡃࡢ⣽⬊ᰴࡢᶞ❧ࡀሗ࿌ࡀࡉࢀ࡚࠸ࡿࠋ

ᚑ᮶ࠊࡇࢀࡽ⣽⬊ࡢᶞ❧◊✲ࡸ⣽⬊ᰴࢆ⏝࠸ࡓᶵ⬟ゎᯒ࡞࡝ࡢศᏊ࣭⏕≀Ꮫⓗ◊✲ࡣࠊ⾑Ύ
ῧຍ᮲௳࡛⾜ࢃࢀ࡚ࡁࡓ (Easty, et al., 1981)ࠋ⾑Ύ୰࡟ࡣࠊ✀ࠎࡢ⺮ⓑࠊ⬡㉁ࡸ⏕⌮άᛶ≀

㉁ࠊࡉࡽ࡟ᮍ▱ࡢᅉᏊࡀᏑᅾࡋ࡚࠾ࡾࠊ⣽⬊ᮏ᮶ࡢᰤ㣴せồᛶࡸቑṪᅉᏊせồᛶࢆṇ☜࡟᳨ウ

ࡍࡿࡇ࡜ࡣ㞴ࡋ࠸ࠋࡉࡽ࡟ࠊ⣽⬊ࡀ⏘⏕࣭ศἪࡍࡿ✀ࠎࡢᅉᏊࢆ⣽⬊ෆศἪᏛⓗ࡟᫂ࡽ࠿࡟ࡍ

ࡿࡇ࡜ࡣᅔ㞴࡛࠶ࡿࠋ
⒴⣽⬊ᰴࡢᶞ❧ࡣ㠀ᖖ࡟ᅔ㞴࡛ (Eun, et al., 2005)ࠊ㛗ᮇᇵ㣴㐣⛬࡛⣽⬊ࡀ⣽⳦ࡸ┿⳦࡞࡝
࡟ởᰁࡋࡓࡾ (Rebecca, et al., 2014)ࠊ⥺⥔ⱆ⣽⬊ࡢ㐣ቑṪࠊࡉࡽ࡟⒴⣽⬊⮬㌟ࡢቑṪ⬟ࡢప
ୗ࡞࡝࡟ࡼࡿࡇ࡜ࡀ୺࡞せᅉ࡜ࡋ࡚ᣲࡆࡽࢀࡿ (Ganjibakhsh, et al., 2017)ࠋ୍᪉ࠊSato ࡽࡣ

ᇵᆅ୰࡟ຍ࠼ࡿ⾑Ύࡢᶵ⬟ࡣࠊ⣽⬊ࡢቑṪ࣭ศ໬࡟࡜ࡗ࡚ᚲせ࡞࣍ࣝࣔࣥࡸ⣽⬊ቑṪᅉᏊࢆ౪
⤥ࡍࡿࡇ࡜࡛࠶ࡿ࡜ࡢ௬ㄝ࡟ᇶ࡙ࡁࠊ↓⾑Ύᇵ㣴ἲࢆ㛤Ⓨࡋࡓ (Hayashi and Sato, 1976;
Barnes and Sato, 1980)ࠋ↓⾑Ύᇵ㣴ἲࡣࠊᇶ♏ᰤ㣴ᇵᆅ࡟࢖ࣥࢫࣜࣥࡸࢺࣛࣥࢫࣇ࢙ࣜࣥ࡞

࡝ࡢ᪤▱ࡢ࣍ࣝࣔࣥࠊ⤖ྜ⺮ⓑࡸ⣽⬊ቑṪᅉᏊ࡞࡝ࢆຍ࠼ࡓ↓⾑Ύᇵᆅࢆ⏝࠸࡚ࠊṇᖖཬࡧ⒴

4

⣽⬊ࡢቑṪ࣭⥔ᣢࢆྍ⬟࡟ࡍࡿ᪉ἲ࡛࠶ࡾࠊ⌧ᅾࡲ࡛࡟⾑⟶ෆ⓶⣽⬊௨እࡢ⣽⬊ࡣ඲࡚↓⾑Ύ
ᇵ㣴ἲ࡛ᇵ㣴ࡀྍ⬟࡛࠶ࡿ (Sato, et al, 2002)ࠋ

㏻ᖖࡢ⏕య࡟࠾࠸࡚⾑Ύ࡜┤᥋᥋ࡋ࡚࠸ࡿ⣽⬊ࡣࠊ⾑⟶ෆ⓶⣽⬊௨እ࡟Ꮡᅾࡋ࡞࠸ࠋࡘࡲ

ࡾࠊ࡯࡜ࢇ࡝ࡢ⣽⬊ࡣ⾑ΎᡂศࢆࡑࡢቑṪ࣭ศ໬ࡢ⥔ᣢ࡟ᚲせ࡜ࡋ࡚࠸࡞࠸ࡇ࡜ࢆព࿡ࡋ࡚࠸

ࡿࠋ౛࠼ࡤࠊཱྀ⭍⢓⭷ࡸ⓶⭵ࡢୖ⓶⤌⧊࣭⣽⬊ࡣࠊ㛫㉁⤌⧊࠿ࡽᇶᗏ⭷ࢆ⤒⏤ࡋ࡚౪⤥ࡉࢀࡿ

ᰤ㣴ᅉᏊ࡛ቑṪ࣭ศ໬ࡀ⥔ᣢࡉࢀ࡚࠸ࡿࠋ⏕య࡛ࡣୖ⓶⣽⬊ࡣ㠀ᖖ࡟άⓎ࡟ቑṪࡋ࡚࠸ࡿࡀࠊ

㛫㉁⤌⧊ࡢ⥺⥔ⱆ⣽⬊ࡣ࡯࡜ࢇ࡝ቑṪࡋ࡚࠸࡞࠸ࠋᚑ᮶ࡢ⾑Ύῧຍᇵ㣴ἲཱྀ࡛⭍⢓⭷⤌⧊ࢆᇵ

㣴ࡍࡿ࡜⥺⥔ⱆ⣽⬊ࡣ㧗࠸ቑṪ⬟ࢆ♧ࡍࡀࠊୖ⓶⣽⬊ࡣ࡯࡜ࢇ࡝ቑṪ࡛ࡁ࡞࠸஦ᐇࡣࠊ⾑Ύῧ

ຍᇵ㣴ἲࡀ⏕యᶵ⬟ࢆ཯ᫎࡋ࡚࠸࡞࠸ࡇ࡜ࢆ♧ࡍࡼ࠸౛࡛࠶ࡿࠋ

↓⾑Ύᇵ㣴ୗ࡛ࡣࠊᇵ㣴ึᮇ࡟ࡣ⥺⥔ⱆ⣽⬊ࡢቑṪࢆㄆࡵࡿࡶࡢࡢࠊ㏿ࡸ࠿࡟ࡑࡢቑṪࢆ೵

Ṇࡍࡿࠋࡇࢀ࡟ᑐࡋࠊ⭘⒆⣽⬊ࡣ↓⾑Ύᇵ㣴ୗ࡟࠾࠸࡚ࡶቑṪࡀ⥔ᣢࡉࢀࡿࡓࡵࠊ⥺⥔ⱆ⣽⬊

ࡢΰධ࡞ࡋ࡟⭘⒆⣽⬊ᰴࡢ༢㞳ࡀᐜ᫆࡜࡞ࡿ฼Ⅼࢆ᭷ࡋ࡚࠸ࡿࠋࡇࡢࡼ࠺࡟↓⾑Ύᇵᆅࢆ⏝࠸

ࡿࡇ࡜࡟ࡼࡾࠊ⾑Ύ࡟ྵࡲࢀࡿᵝࠎ࡞⺮ⓑ➼ࡢᙳ㡪࡞ࡋ࡟ࠊ⭘⒆⣽⬊ࡢቑṪ࣭ศ໬➼ࡢヲ⣽࡞

࣓࢝ࢽࢬ࣒ࡢゎᯒࡸࠊ✀ࠎࡢ⏕⌮άᛶศᏊࡸᢠ⒴๣ࡢస⏝ᶵᗎࢆ᳨ウࡍࡿࡇ࡜ࡀྍ⬟࡜࡞ࡿࠋ

ⴭ⪅ࡢᡤᒓࡍࡿ◊✲ᐊ࡛ࡣࠊཱྀ⭍⒴ࡢ⏕᳨⤌⧊ࡸᡭ⾡ษ㝖⤌⧊ࡼࡾࠊ↓⾑Ύᇵ㣴ἲࢆ⏝࠸࡚⒴
⣽⬊ᰴࡢᶞ❧◊✲ࢆ⾜࠸ࠊࡇࢀࡽ⣽⬊ᰴࡀⓎ⌧࣭⏘⏕ࡍࡿ⣽⬊ቑṪᅉᏊࠊཷᐜయཬࡧ exosome

࡞࡝ࡢゎᯒࢆ㏻ࡋ࡚ࠊཱྀ⭍⒴⣽⬊ࡢ≉ᛶࢆ⣽⬊ෆศἪᏛⓗ࡟᫂ࡽ࠿࡟ࡋ࡚ࡁࡓࠋ

5

ᮏ◊✲࡛ࡣࠊཱྀ⭍ᡥᖹୖ⓶⒴ཬࡧཱྀ⭍㡿ᇦ࡬ࡢⓎ⏕ࡀ⛥࡛ࠊ⥅௦⥔ᣢྍ⬟࡞⣽⬊ᰴࡢᶞ❧ࡢሗ

࿌ࡢ࡞࠸ཱྀ⭍⭢ᡥᖹୖ⓶⒴ࠊࡉࡽ࡟↓⾑Ύ᮲௳ୗ࡛ࡢᇵ㣴ᡤぢࡸච␿⤌⧊Ꮫⓗ᳨⣴ࡢ⤖ᯝࢆ⑓

⌮㒊࡟᝟ሗᥦ౪ࡍࡿࡇ࡜࡛☜ᐃデ᩿࡟⮳ࡗࡓࠊྠࡌࡃཱྀ⭍㡿ᇦ࡬ࡢⓎ⏕ࡀᴟࡵ࡚⛥࡛࠶ࡿཱྀ⭍

ཎⓎ⚄⤒ෆศἪ⒴ࡢ⣽⬊ᰴࡢᶞ❧◊✲ࢆࠊ↓⾑Ύᇵ㣴⣔ࢆ⏝࠸࡚⾜ࡗࡓࠋࡉࡽ࡟ࡇࢀࡽ⣽⬊ࡢቑ

ṪᅉᏊせồᛶࡸ㑇ఏᏊⓎ⌧࡞࡝ࡢ≉ᛶࢆࠊ⣽⬊࣭ศᏊ⏕≀Ꮫⓗ࡟᫂ࡽ࠿࡟ࡍࡿࡇ࡜ࢆ┠ᣦࡋࡓࠋ

6

➨ 2 ❶  ཱྀ⭍⒴⣽⬊ᰴࡢᶞ❧
➨ 1 ⠇ ᐇ㦂ᮦᩱཬࡧ᪉ἲ
(1) ᇵ㣴ᮦᩱཬࡧᇵ㣴᪉ἲ

ⴭ⪅ࡢᡤᒓࡍࡿ◊✲ᐊ࣭デ⒪⛉࡛἞⒪ࢆ⾜ࡗࡓࠊཱྀ⭍ᡥᖹୖ⓶⒴ࠊཱྀ⭍⭢ᡥᖹୖ⓶⒴ࠊཱྀ⭍ཎ
Ⓨ⚄⤒ෆศἪ⒴ᝈ⪅ࡢ⭘⒆⤌⧊ࢆ᥇ྲྀᚋࠊ㏿ࡸ࠿࡟ึ௦ᇵ㣴ࢆ⾜ࡗࡓ (ᗈᓥ኱Ꮫࣄࢺࢤࣀ࣒೔⌮
ᑂᰝጤဨ఍ᢎㄆィ⏬◊✲➨ࣄ 58 ྕཬࡧ 191 ྕ)ࠋ
ᇶ♏ᰤ㣴ᇵᆅ࡜ࡋ࡚ࠊDulbecco’s Modified Eagle’s Medium (Dulbecco and Freeman,
1959)(Sigma-Aldrich, St. ...

この論文で使われている画像