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Title
ハウスダストマイトによって活性化されたイヌケラチノサ
イトにおけるGM-CSFの産生機序に関する研究( 本文
(Fulltext) )
Author(s)
木村, 剛
Report No.(Doctoral
Degree)
博士(獣医学) 甲第412号
Issue Date
2014-03-13
Type
博士論文
Version
ETD
URL
http://repository.lib.gifu-u.ac.jp/handle/123456789/49035
※この資料の著作権は、各資料の著者・学協会・出版社等に帰属します。
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AD: Atopic dermatitis̖DeqˆŲȪʀǻ̗
AP-1: Activator protein-1
AQP: Aquaporin
CAD: Canine atopic dermatitis̖FiDeqˆŲȪʀǻ̗
CCL: CC chemokine ligand
CCR: CC chemokine receptor
CsA: Ciclosporin A
Der f: Dermatophagoides farinae̖TgoGo_h̗
Der p: Dermatophagoides pteronyssinus̖RoGo_h̗
DPBS: Dulbecco’s phosphate-buffered saline
E-64: Trans-Epoxysuccinyl-L-Leucylamido-̖4-Guanidino̗Butane
ERK: Extracellular signal-regulated kinase
GAPDH: Glyceraldehyde 3- phosphate dehydrogenase
GM-CSF: Granulocyte macrophage colony-stimulating factor
HDM: House dust mite̖lGY_YeyFe̗
Ig: Immunoglobulin̖ÅȝQ†sƒ‡̗
IL: Interleukin
INPPL: Inositol polyphosphate phosphatase-like 1
LPS: Lipopolysaccharide̖ƒxħɓ̗
MHC: Major histocompatibility complex̖“ʘɜɧ˕ûƅðʖû­̗
MMP: Matrix metalloprotease̖ɚɹĦĚʶÔʞ˛ɖ̗
NFAT: Nuclear factor of activated T-cells̖ǥŲç T ɚɹNjËĎį̗
5
NF-κB: Nuclear factor-kappa B
NK: Natural killer
NLR: Nucleotide binding oligomerization domain-like receptor̖NOD ǒöĺ
­̗
PAR: Protease activated receptor̖t†cDˆ\ǥŲçöĺ­̗
PCNA: Proliferation cell nuclear antigen
PMA: Phorbol 12-myristate 13-acetate
RTPCR̠Reverse transcription polymerase chain reaction
SAA: Serum amyloid A
SGPL: Sphingosine-1-phosphate lyase
SPINK: Serine protease inhibitor Kazal
SPL: Sphingosine 1-phosphate lyase
S1P: Sphingosine 1-phosphate
TEWL: Transepidermal water loss̖ɝʓȪǟÔʍƝˠ̗
TGF: Transforming growth factor
Th: Helper T
TIMP: Tissue inhibitor of metalloproteinase
TLR: Toll like receptor̖Toll ǒöĺ­̗
TNF: Tumor necrosis factor
TRIF: Toll/IL-1 receptor/resistance domain-containing adaptor-inducing
IFN-β
TSLP: Thymic stromal lymphopoietin
6
ɥʫ
7
FiDeqˆŲȪʀǻ̖Canine atopic dermatitis, CAD̗.ǻȡŲȢŲ
-D…„OˆŲȪʀȟŴ(=˗¨Ȩ+ɖĎ˫'=Immunoglobrin
̖Ig̗E ƅ­,˫ˋ!ȅūȨ+ʃřȡȇBĀ>ȟŴ(>)ĵɭ?'
>[40]ȅūȨ+ʃřȡȇ).ȪʀɑʁĠș˙(>ȲĂĐ÷Ĉ;0ɲ˧
ĂĐ-3ɽɅ;0Ëɳ)$!˪ļ˙ɱÛ˿;0ʺ˪-ȫ:˱Ò
ţž?>˙«'ɿ˙ɰŸÛ˿;0Ħɰˑ,ıɍȨ+Ȣʉ̔
<óũ'ʦ8<?>)(>CAD ,'.ȡ±-ɕ 70̕lGY_
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)<HDM ȠŹ,'˞ʘ+ťàBDŽ!'>)űA?>[46 75]
+A#HDM .­Ë,´Æ!Ŧɴdzɚɹ,ɞû! HDM ȅȜȨ IgE
BdžǕ>),;$'ɴdzɚɹ-ǥŲçBŠʹǥŲç?!ɴdzɚ
ɹ<.ǻȡŲ|dEIˆ^ˆƚÓ?îƮė-ːƛȡB“­)>ǻȡ
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t?'!
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Ƥȓ Ȇ,'.HDM -ɝȪȨƱ˽,;$' CAD ǒ-ʃřȡȇȦ
Ȏ>)Ƭ<)+$'>[95]<,CAD ȡ±,'.Ȫʀm
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'>+A#CAD -ȠŹ,'.ƅð)ʓȪBǑž>ɚɹɬ)P†YeˆP˞ʘ+ťàBDŽ!'>øɺŲ>)>
R‚`kVFe.ʓȪ-ɕ 95%Bí8>ɚɹ(>?5(ȄȐȨ+
m ƒ D ) ' - ǖ ɺ - 6 ǣ Ȯ ? ' ! CC chemokine ligand 17
8
(CCL17)[123]Thymic stromal lymphopoietin (TSLP)[13];0 Granulocyte
macrophage colony-stimulating factor (GM-CSF)[92]+*-VFeLF‡9Ȕ
ȓ>)oe;0yGY,'ȸ?!Fi-R‚`kVFe,
'.TgoGo_h̖Dermatophagoides farinaeDer f̗-“ʘD…„
S‡(> Der f 1 -ÚǸ,;$'
Interleukin
̖IL̗
-8GM-CSF ;0 Tumor
necrosis factor̖TNF̗-α - mRNA ʽÍˠġã>)Ƭ<)+$'
>[72]-;,R‚`kVFe,.ǒ+VFeLF‡BȔȓ>ɺâ
>CAD -ȠŹ,>úĎį-ťà,%'.Ƭ<,+$'+
(ƼȵɄ(.HDM -ÚǸ,;$'R‚`kVFe<Ȕȓ?>ǪŲ
Ďį-#CAD -ȠŹ,˫>ÔįBüĵ-ȔȓʧŁ-|LhZ{
BƬ<,>)BȮȨ)'ȵɄBʒ$!
9
ɉ1ɇ
FiR‚`kVFe,'
lGY_YeyFe,;=ʽÍʧŁ?>
˗¨įɬ-ɢɪȨʞǃ
10
1 .1. Śʫ
CAD .˗¨ȨɷƯBƶ> IgE ŸēŲ-D…„OˆŲǻȡŲȢŲȪʀȟŴ
(>[40]­Ë,´Æ!ƅð,;$'ǥŲç?! B ɚɹ.u„nˆ̣
ɚɹ<Ȕȓ?! IL-4 5!. IL-13 -İē,'IgE BȔȓ>ţʶ
ɚɹ1)Ôç>[35]?5(CAD ,>ƅð-Ʊ˽.ăÿČ;0
ǩçɌ-ɑʁBŸ'>9-)ɮ<?'![89]+<Ƥȓ Ȇ
,Ŀ> HDM -ɝȪȨƱ˽,;$' HDM ȅȜȨ IgE -Ȕȓµˍ?
<,Ȫʀ1- HDM -ÌƱ˽,;$' CAD ǒ-ȪʀȠĥȓ>)Ƭ<
)+=[95]Ȫʀ. CAD ,>ƅð-˞ʘ+Ʊ˽ɝʻ(>)ȸ?!
Ɣ̈-Ȫʀɜɧ-ƵĦŇ,.ʓȪİē'>ʓȪ.R‚`kVFe
,;$'Ǒž?ţŹIJȨ,̆ɐŇƶǎŇ;0ĚśŇ,Ô̈>)
(>ʝŇ.̆ɐŇ-ɚɹɼNj!ŦɚɹʶË-žÔBĚ,ţž?>
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0yGY,'.Ôįˠ 500 _„e‡¤Œ-Ȅʶ.ˉː+)ɮ<?'
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àBDŽ!'>ȪʀmƒDǖɺ-ʣ³Ǣ)'ɝȪȨǟÔʍƝˠ
̖Transepidermal water loss, TEWL̗-DZĵȴ<?'>[94]TEWL .
Ȫʀ<ʍȦ>ǟÔ-àûBȸ'=Ȫʀ-ˈːŲ;0mƒDǖɺB
ʣ³>ƂǢ-̚%(>CAD ȡ±(. TEWL ̒¼Bȸ!)<
Ȫʀ-mƒDǖɺ¬'>øɺŲȸ?![23]<,CAD ȡ±,
'.ʝŇɻʶ(>[‚zf-ǯł) TEWL -ŒƫȰ˫'!)
<[‚zfmƒDǖɺ,˫'>)ɮ<?>[32]5!CAD Ķ̐Ȇ,'ʝç,'˞ʘ+ťàBDŽ!'>rE‚Qƒ‡½
œȆ,<2'ǯł'>)ěā?'>[74]<,CAD }d„
11
Ȇ-Ƞĥ˙(.½œȆ(Ė‰,ȦȎ'>.-rE‚Qƒ‡ơȇ%
ŽĖ‰+ȦȎn^ˆ‡Bȸ!)ěā?'>[104]
mƒDǖɺBɡƈ>Œ(˞ʘ+[‚zf;0rE‚Qƒ‡.R‚`kV
Fe,;=Ȕȓ?>)oe,'ȸ?![6][‚zf.R‚`kV
Fe-ʝç-ːȿ(Ȕȓ?>ʝŇɻʶ(=ƶǎŇ;0̆ɐŇ([ƒ‡
)n„zeF„-CoA <ûž?-ŦQƒTW„[‚zf5!.YrE
‡UzIƒ‡)'ʎɃ?ʝŇɚɹ˪(Q„T[…t†W^ˆ\5!.Y
rE‡UzIƒgˆ\,;=Ìûž?>[48, 80]5!rE‚Qƒ‡.̆ɐ
ŇR‚`kVFe,'ɚɹ̑nj)+>ɚɹʶË-R‚`‡ɦɡBɞû
Ð˹!ŦʝҌŇ(.LYnˆ\ 14 +*,;$'Ôʞ?ĩǾ·DzĎ
į)'Á^‡nP(>)ɮ<?'>[18]rE‚Qƒ‡.̆ɐŇR‚
`kVFe-ƈ%R‚eoDƒ‡̆ɐË,'Û̏­-t†rE‚Qƒ‡)
'ȓûž?wtX`„D„Oh‡dFzgˆ\,;$'ãǟÔʞ?'
rE‚Qƒ‡)+>)oe,'ěā?'>[18]mƒDǖɺ,˫
>ÔįR‚`kVFe,;$'Ȕȓ?>)B˥6>)R‚`kVF
e-ǖɺȜœ. CAD -ȠŹ,'˞ʘ+ťàBDŽ!'>)ɮ<?>
ʓȪ- 95̕BǑž>R‚`kVFe.Ħ˙<-ÚǸ,;$'ǥŲç
?ǒ+ǪŲĎįBȔȓ>CAD ȡ±-Ƞĥ˙,'.ʂȪȡ-“+
ðĎʊ(> Staphylococcus pseudointermedius -R‚`kVFe1-¢ȳ
ŲġŢ'>)ȸ?'>[113]5!CAD -źŲȠĥ(.Ȣ6
,;>ʄ¿:ʓȪ-ɻʶȜœ,;$'ʓȪȑĠȶɣɚʊT†hˆţž
?'>Ĝûħ[26]ɚʊ-ʊ­žÔ(>ƒxħɓ̖Lipopolysuccaride,
LPS̗,ÚǸ?!Fi-ę̍R‚`kVFe,'.t†Y^Q‚‡dE
‡ E2 Ȕȓ?>[7]<,HDM ,;$'ǥŲç?!FiR‚`kVF
e-ɚɹNJ,'.ĭ’ȏ-ˏʸ,˫ˋ> IL-8 ;0̆ɐȏɔ:ìȏɔ
12
-ǥŲçĎį(> GM-CSF - mRNA ʽÍˠ-ġãƬ<)+$'>
[72]?<-ɞDŽ<ʓȪ.ȄȐȨmƒD-6+<R‚`kVFe
<Ȕȓ?!VFeLF‡BŸ!ÅȝȨmƒDǖɺ-ťà9DŽ!'>)
űA?>
Der f ;0 Der p .ȑĠ’,İē> HDM -“ʘ+D…„S‡(>[64]
CAD ȡ±- 54.4%,'Der f ;0 Der p ,Ŀ>ʑǮ’-ȅȜȨ IgE
¼-ġãěā?'>[75]ȅ, Der f ,Ŀ'.ȪËóůʤ̐,'
9ȡ±- 64.3 %(˳Ųóůʦ8<?'>[75]Der f ;0 Der p .
?
?ɕ 20 ɀ̈-D…„S‡^‡nPBþƶ?<-D…„S‡-’(9
ȅ, Der f 1 ;0 Der p 1 . IgE )-ɞûȍ̒)oe;0yGY,
'ȸ?'>[45, 50, 110]Der f 1 ;0 Der p 1 .t†cDˆ\(
=?<-t†cDˆ\ǥŲ,;>ŒȪɚɹ˪ɞû-ȶĢ:ɚɹʓ˿-Ôį
-Õƣɚɹ-ǥŲçBŠʹ[45]Der f 1 ;0 Der p 1 .ŒȪɚɹɚɹʁŒ,ȦȎ>t†cDˆ\ǥŲçöĺ­̖Protease activated receptor,
PAR̗-2 BŸ'oeR‚`kVFeBǥŲçIL-8 ;0 GM-CSF BȔ
ȓ>[60]5!Der p 1 PAR-2 WQg„ɝʻ).Ø,ɚɹËn^ˆ
‡ʦʭöĺ­(> NOD ǒöĺ­̖Nucleotide binding oligomerization
domain-like receptor, NLR̗rCzƒˆ-̚%(> NLRP3 BŸ'oeR
‚`kVFeBǥŲçIL-1β ;0 IL-18 -ȔȓBʧŁ!)ěā
?'>[25]?<-ɞDŽ.ȓ­Ë,´Æ!D…„S‡. I ėːƛȡBŠ
ʹ/=(+ŒȪɚɹ)-Ȱ˜¯ȕBŸ!Ȝœ+ɚɹŲÅȝBʧ
Ł'>øɺŲBȸć>9-(>CAD -ȠŹ1-˫.Ƭȷ(.+
!$'ɉ‰ɇ(.CAD -ȠŹ,>t†cDˆ\ǥŲBƶ>
D…„S‡)ŒȪɚɹ-Ȱ˜¯ȕBƬ<,>)BȮȨ)'HDM ,;
$'ǥŲç?!FiR‚`kVFeɚɹNJ,>˗¨į-ʽÍʧŁB
13
DNA yFP†D…F,;$'ɢɪȨ,ʞǃ!
14
1 .2. ƾƢ)ƥǢ
1.2.1. ƾƢ
ĚśŇR‚`kVFe)'-ŲȇBƈ%)ƨ,ȸ?'=[112]F
i-ǻȡŲȪʀȟŴ-Ķ̐bˆ„)'-ƶȕŲȸ?'> CPEK[72,
112] ;0-ę̍Ǫ(> CnT-09 . CELLnTEC Advanced cell systems
̖Bern, Switzerland̗;=ÆƂ!ǽʑǮęĕ(> Opti-MEM I ;0ɠ
£ę̍Ʈ-ɚɹÝ˺-˶,ȕ!eƒtW‡ÑȐÞ̖TrypLE Express̗.
Invitrogen̖Carlsbad, CA̗<ÆƂ!Dulbecco’s phosphate-buffered
saline̖DPBS̗. Sigma-Aldrich̖St. Louis, MO̗-9-B°ȕ!Der f
-“ʘD…„S‡(> Der f 1 -ɒʕȄ.ȓçIJmFKpXjYNJş§ȹ̖ǂ
œ̗;=ʷÆ!
1.2.2. ɚɹę̍
̞č< 10 čɠ£! CPEK B 10 mL - CnT-09 ,Ǩˏ375% CO2
-ȑĠ( 75 cm2 r‚YT̖NUNC, Roskilde, Denmark̗Bȕ'ę̍!
80%T‡r„I‡e,˒!Ǜ˴(ę̍ǪB˰ñ10 mL - DPBS (ɚɹ
BǤǧ!TrypLE Express B 3 mL ã!Ŧ37,' 5-10 Ô˪˾ɨ
ɚɹBÝ˺!CnT-09 B 7 mL ãɕ 80̕-ɚɹÝ˺'Ǩˏ!
)Bȷʦ!Ŧ10 mL -ɚɹǨˏǪB 15 mL centrifuge tube̖NUNC̗,
čôķǰ( 180 G5 Ô˪˔Ŭ!˔ŬŦŒǮB˰ñɚɹĝB CnT-09
,;$'˾,ÌŽǹ!ÌŽǹ!ɚɹǨˏǪB2×104 /mL -ĻŜ(
24-well plates̖NUNC̗, 1 mL %Ôǣ80̕T‡r„I‡e,˒>5
-Ŧ
ę̍ǪB Opti-MEM I ,ĥƳ
( 37 5%CO2 -ȑĠ(ę̍!
15
ǽʑǮ( 24 Ʈ˪ę̍!24 Ʈ˪Ŧ, Der f 1̖10 μg/mL̗Bǭã!
Opti-MEM I 5!. Opti-MEM I -6,ƴ'37 5%CO2 ( 24 9.
48 Ʈ˪ę̍!
1 .2.3. CPEK <-eˆ^„ RNA ƆÓ
ę̍ɚɹ< RNeasy Mini Kit̖Qiagen, Valencia, CA̗B°ȕ'eˆ^
„ RNA BƆÓ!čô!eˆ^„ RNA .TURBO DNA-Free Kit
̖Applied Biosystems, Foster City, CA̗B°ȕ'Sk{ DNA -˰ñBʒ$
!Ŧ-80(·İ!
1.2.4. yFP†D…F
Canine KƒU DNA yFP†D…F Ver.2.0
̖4×44K̗
̖Agilent Technologies,
Palo Alto, CA̗Bȕ!ʞǃBNJş§ȹmFKyeƒaPYȵɄƁ̖êʌ̗,
'ĶƦ?Ƽʞǃ.¤-Ƃ̄(ʒA?!Quick Amp Labeling Kit̖Agilent
Technologies̗Bȕ'úV‡t„-eˆ^„ RNA < 2 Ƽˣ cDNA Bû
žin vitro ʽÍ,;= cRNA Bûž!-ƮCyanine ʆɖ(Ǔʭ
?! Cyanine 3-CTP Bõ=˃5ʐÄǓʭBʒ$!Gene Expression
Hybridization Kit̖Agilent Technologies̗B°ȕǓʭ cRNA BlFsƒ
_F\ˆW‡marCˆ,ã
Canine KƒU DNA yFP†D…F Ver 2.0
̖4×44 K̗Œ( 17 Ʈ˪yFP†D…FŒ- DNA )óů!
ǤǧŦAgilent
Microarray Scanner ( DNA yFP†D…F-F|ˆXBʪ6õ=Feature
Extraction Software̖v.9.5.3.1̗Bȕ'úYxae-ʐÄWQg„Bƞ¼ç
!
16
1 .2.5. ʞǃƥǢ
Der f 1 -ǭã 24 Ʈ˪Ŧ;0 48 Ʈ˪Ŧ,'HDM ǭãɬ,>ʽ
Íˠǽǭãɬ)ǜʾ'2 º¤Œ-ȦȎġã5!. 1/2 ¤-ȦȎ¬Bȸ
!˗¨įƞ-ĥæBʣ³!<,oe- Atopic dermatitis̖AD̗Ƞ
ĥ˙,'ȦȎ-ĥçʦ8<?'> 52 ˗¨į[53, 85, 114]̖ʓ 1-1̗CAD
Ƞĥ˙,'ȦȎĥçʦ8<?'> 33 ˗¨į[77, 126]̖ʓ 1-2̗;0
R‚`kVFe(ȦȎ>ʄǾÅȝ;0ȌŧÅȝ,˫ˋ>VFeLF‡22 ˗¨į[1, 21, 37]̖ʓ 1-3̗-ĥæ,%'9ʣ³!
1.2.6. ĵˠƒD„^F{ RT-PCR Bȕ!Ǐʢ
yFP†D…F-ʞǃɞDŽ.1.2.5 ,;$'˖Ó!˗¨įBȅȜȨ,ġŔ
>t‚Fyˆ̖ʓ 1-4̗Bȕ!ĵˠƒD„^F{ Reverse transcriptional
polymerase chain reaction̖RT-PCR̗,;$'Ǐʢ!ĵˠƒD„^F{
RT-PCR .yFP†D…Fʞǃ(ȕ!eˆ^„ RNA Bˡė)! 1 Ycat
RT-PCR̖One-Step SYBR PrimeScript RT-PCR Kit II, ^L‚mFK, Ƕʳ̗
,;$'ʒ$!2'-óů. Thermal Cycle Dice Real Time System̖^
L‚mFK̗Bȕ'ʒˇʽÍóůB 42 5 Ô˪;0 95 10 Ƚ˪
xƒ|‚ˆ\ˋˣóůB 95 5 Ƚ˪;0 60 30 Ƚ˪B 40 VFP„ʞ˺
. 95 15 Ƚ˪60 30 Ƚ˪95 15 Ƚ˪-ǀ¥(ĶƦ!ȮȨ˗¨į
-ʽÍˠ. Glyceraldehyde 3-phosphate dehydrogenase̖GAPDH̗BƒrC
…‡Y˗¨į)!ȰĿĵˠ¼)'ɋÓ!ŧ<?!ȰĿĵˠ¼B9),
ΔΔCt Ǣ,;$' HDM ǭãɬ)ǽǭãɬ,Ŀ>ȰĿĵˠ¼BɋÓ!¼2 º¤Œ-ʽÍˠġã5!. 1/2 ¤-ʽÍˠ¬($!ĜûB?
?ƶŷ+ġã5!.¬)!
17
1 .3 ɞDŽ
1.3.1 CPEK ,'ȦȎġã;0ȦȎ¬Bȸ!˗¨įƞ-ĥç
̖ʓ 1-5̗
ƅðİē(ę̍! CPEK ,'ɕ 22,000 ˗¨į̖t†ˆs̗-Ȧ
Ȏȷʦ?!ƅð-ǭã,;$'ȦȎĥæ!˗¨įƞ-ɞDŽBʓ 1-5
,ȸȦȎġã;0ȦȎ¬Bȸ!˗¨įƞ.ę̍Ʈ˪)É,ġã>
Àý,$!
1.3.2 oe AD Ƞĥ˙,'ȦȎˠĥç'>˗¨įɬ,%
'-ʞǃ̖ʓ 1-6̗
ːñ,ĶƦ?!oe AD Ƞĥ˙-ɢɪȨʞǃ,'ȦȎĥçěā
?'> 52 ¹-˗¨į,%'ʣ³!
52 ˗¨į-#FiȕKƒU DNA
yFP†D…FNae,;$' 27 ˗¨į-ʞǃøɺ($!- 27 ˗¨
į-#ƅð-ǭã,;$' 20 ˗¨į-ȦȎȷʦ?!ȦȎĥæBȸ
!˗¨į-#AQPAquaporin3IL-8CD26/DPPDipeptidylpeptidase
4 ;0 Keratin10 ,%'. 2 º¤Œ-ȦȎġãʦ8<? CCL17
Keratin5S100A2 ;0 S100A8 ,'. 1/2 º¤-ȦȎ¬ʦ8<
?!Keratin5 ;0 Keratin10 B˰yFP†D…F(ŧ<?!ɞDŽBĵ
ˠƒD„^F{ RT-PCR Bȕ'Ǐʢ!)@AQP3IL-8 ;0 S100A8
,%'.ƅðÚǸ 24 Ʈ˪Ŧ,ʽÍˠ-ġãʦ8<? CCL17 ;0
S100A2 ,˫'.ƅðÚǸ 48 Ʈ˪Ŧ,ʽÍˠ¬!CD26/DPP4 ,%
'.ʽÍˠ-ĥç.ʦ8<?+$!
18
1 .3.3 CAD Ƞĥ˙,'ȦȎĥç'>˗¨įɬ,%'-ʞǃ
̖ʓ 1-7̗
yFP†D…FBȕ! CAD -Ƞĥ˙ʞǃ,'ːñ,ȦȎĥçěā
?'> 33 ¹-˗¨į,%'ʣ³!33 ˗¨į-#FiȕKƒU
DNA yFP†D…FNae(ʞǃøɺ($!˗¨į. 23 ˗¨į($!
-#ƅð-ǭã,;$' 20 ˗¨į-ȦȎȷʦ?!ȦȎȷʦ?
!˗¨į-#FLIPL1Down-regulated in ovarian cancer 1 isoform 2
INPPLInositol polyphosphate phosphatase-like1SAASerum amyloid
A1CD26/DPP4SPINKSerine protease inhibitor Kazal5 ;0 SGPL
Sphingosine-1-phosphatase lyase1 ,%'. 2 º¤Œ-ȦȎġã
AKAP9A-kinase anchor protein 9;0 S100A8 ,%'. 1/2 º¤ȦȎ¬ʦ8<?!ĵˠƒD„^F{ RT-PCR Bȕ!Ǐʢ(.S100A8
̖24 Ʈ˪Ŧ̗TIMP1̖24 Ʈ˪Ŧ̗SPINK5̖48 Ʈ˪Ŧ̗;0 SGPL1̖24
Ʈ˪Ŧ̗,'ʽÍˠ-ġãʦ8<?!SPINK5̖24 Ʈ˪Ŧ̗-ʽÍˠ
-ĥæ.-¡-˗¨į)Ȝ+=24 Ʈ˪Ŧ,¬!Ŧ48 Ʈ˪Ŧ,ǽǭã
Ʈ- 172.02 º,5(ġã!)Ƭ<)+$!
+SAA1 ,'.
ǏÓ?DPP4 ,'.ʽÍˠ-ĥç.ʦ8<?+$!
1.3.4 R‚`kVFe,'ȦȎ>ʄǾÅȝ;0ȌŧÅȝ,˫ˋ
>VFeLF‡̖ʓ 1-8̗
R‚`kVFe,''ȦȎěā?'> 22 ˗¨į-#yFP
†D…F,'ʞǃøɺ($!˗¨į. 19 ¹($!- 19 ˗¨į#12 ˗¨į,'ƅðÚǸŦ,ȦȎʦ8<?!ȦȎʦ8<?!˗
¨į-# IL-8IL-15CCL2CCL20TGF-2 ;0 GM-CSF . 2 º¤
Œ-ȦȎġãBȸ!CCL17̖48 Ʈ˪Ŧ̗. 1/2 º¤-ȦȎ¬Bȸ
19
!ĵˠƒD„^F{ RT-PCR ,;$'Ǐʢ!ɞDŽIL-8CCL20GM-CSF
;0 IL-15 ,%'.ƅðÚǸ 24 Ʈ˪Ŧ,5!TGFTransforming
growth factor-2 ,%'.ƅðÚǸ 48 Ʈ˪Ŧ,ʽÍˠġã!ȅ,
IL-8CCL20 ;0 GM-CSF ,'.ƅðÚǸ 48 Ʈ˪Ŧ, 8 º¤Œ9-ʽ
ÍˠġãƬ<)+$!
20
1 .4 ɮļ
R‚`kVFe.ʓȪ,>ȄȐȨçIJȨ;0ȓȄIJȨmƒDBǑ
ž>˞ʘ+ɚɹžÔ(>R‚`kVFe,>˗¨į-ʽÍˠ.əĦ
ɦ:çIJȄʶ+*ǒ+ÚǸ,;$'ĥæ>)oe:yGY,'
ȸ?'>[39, 93, 98]yFP†D…FBȕ!ʞǃ(.N-D[`„-LWYcF‡-ǭã,;$' 334 ¹-˗¨įoɖǭã,;$'. 121 ¹-˗¨
į-ȦȎĥæoe,'ʦ8<?![39, 93]žč-ȵɄ(.24 Ʈ˪Ŧ,
1,344 ¹-˗¨į<,ę̍ 48 Ʈ˪Ŧ,. 3,748 ¹-˗¨į-ȦȎĥæʦ
8<?!)<Der f 1 ,.N-D[`„-L-WYcF‡:oɖ+*BŒč>
R‚`kVFe,Ŀ>Ţâ+ʽÍʧŁ5!.ǥŲƃÙİē>)ȸ
?!HDM -ƅð^‡nP(> Der f 1 .ńË-Ę 1 g ’,'.ŕ
Ė 2.454.97 g İē'=ħ)@(. 30 g ¤Œİē>Ĝû9
>[99]5!CAD ɩŴFiB̌ɵ>ńË-Ę,İē> Der f 1 .̌
ɵFi CAD ,ɩŴ'+Ĺŝ)ǜ2'ħ)ȸ?'>)
<[31]ƼȵɄ(ȕ! Der f 1 ǺŜ̖10 g/mL̗.Ķ˶-ȑĠ’)34ü
ȿŜ-ǺŜ(>)ɮ<?>
ɢɪȨ+˗¨įȦȎʞǃ,;$'?5(ȠŹ1-˫ŽƬ($!Ô
įƬ<)+>Ĝû>ʞǃƥǢ)'ȦȎn^ˆ‡-̈ªŲ,Ě&
ɔɟǔB¯ž>˴ŇȨP‚Y^ƒ‡QƵ9;°ȕ?'=ȦȎĥ
ç-n^ˆ‡<-ǖɺB–DZ(>øɺŲ>[122]5!ƆÓ?
!˗¨į,'.ƨȴ-ŵěBĚ,˗¨į-ʽÍ,!>ɝʻB–DZ
>)øɺ)+$'>[122]oeyGY;0‚ae-ŵě,Ě&'
˚ÖȰüŲ˗¨į-ȦȎn^ˆ‡BɢɪȨ,Əɘ>)Fi,'9ø
ɺ(>žč-ȵɄ,'.?<-ʞǃBʒ!8-DtƒRˆW
21
‡]reGHD°ȕ(+$!(ƼȵɄ(.?5(,ěā$!oe;0Fi- AD Ƞĥ˙-yFP†D…Fʞǃ,'ȦȎĥçʦ8<?!˗¨įɬ;0R‚`kVFe,>ȦȎʦ8<?>Åȝ˫ˋ
Ôįɬ,ȳȮʞǃBʒ$!
oe- AD Ƞĥ˙,'ȦȎġŢʦ8<?'> AQP3 .ǟ;0Qƒ
[†ˆ„-ˀˆ­(> Aquaporin3 BTˆf>˗¨į(>[41, 71]
AQP3
BkaPDGe!yGY(.ʓȪɜɧ,>ǟ;0Qƒ[†ˆ„-ˈ
ːŲ.?9¬>)ȸ?'>[71]5!uD…YyGY-˟ȓ
ė) AQP3 ǗƖė,>ʓȪ,þƶ?>ǟÔˠBǜʾ!)@AQP3
ǗƖėuD…YyGY(.˟ȓėyGY;=9¬'>)Ƭ<)+
$'>[71]<,AQP3 kaPDGeyGY,Ŀ'D…„S‡(>
ovalbumin BɝȪȨ,ĞŐ'9ʓȪ-ɴï.ʦ8<?ɚɹġǚ-ƉǓ)
+> PCNA̖Proliferation Cell Nuclear Antigen̗˳Ų-R‚`kVFe.˟
ȓė,ǜ2'ƶŷ,ł+)ȸ?'>[81]ltc‡(> Oxazolone
BĞŐ! AQP3 kaPDGeyGY,'.ʑ’- CCL17 ;0ɤ IgE
˟ȓėyGY,ǜ2'ƶŷ,ġã'!)<AQP3 .ʓȪ-mƒD
ǖɺ,˫>)Ƭ<)+$![81]Fi,>ʥɚ+Ǐʡ.ʒA?'
+oe;0yGY,>?5(-ěā)žčŧ<?!ɞDŽ<
HDM ,;>ÚǸ.R‚`kVFe,> AQP3 mRNA -ʽÍˠBġã
>),=ʓȪǟÔþƶˠ-ɡƈ)ʓȪɴïBŸ'mƒDǖɺ-čũBʧ
Ł'>øɺŲɮ<?!
Fi;0oe- AD Ƞĥ˙(-ȦȎġŢʦ8<?'> SGPL1 .
Sphingosine 1-phosphate lyase̖SPL̗BTˆf>˗¨į(>[108, 126]
SPL . ʓ Ȫ m ƒ D ɻ ʶ ( > [ ‚ z f - £ ʬ Ȕ Ȅ ( > Sphingosine
1-phosphate̖S1P̗BÔʞ>[109]S1P .DxeˆWYBƃÙ>)(
22
ɚɹġǚ;0ÔçBµˍ>)ȴ<?>[65]‰ƥSPL ˗¨įkaP
DGe F9 ɚɹBȕ!ȵɄ,'S1P ʄʼ(.+SPL ɚɹ-£ʬ
BšˍġǚBµˍ>)ȸ?![57]oeR‚`kVFe,'9
SPL ÔçB˪ƐȨ,µˍ>)ȸ?![11, 44]CAD Ƞĥ˙,>
S1P -ȦȎ¬'>)BɮŻ>)[8]HDM -ɝȪȨ+ÚǸR‚
`kVFe,> SGPL1 -ʽÍBʧŁR‚`kVFe-ġǚ;0Ôç
Bµˍ'>øɺŲȸ?!
Serine protease inhibitor Kazal-typeSPINK5 .Fi;0oe- AD
Ƞĥ˙,'ȦȎġŢ>)ȸ?'>[84, 126]SPINK5 BkaP
DGe!yGY-Ȫʀ,'.̓Ņţž)ʝŇÝ˺-šˍ;0ǝ-ţ
žȜœʦ8<?![27]<,̆ɐŇ-ƐȳÔį(>dY}Q…F‡̚ȦȎ¬-6+<˼į̇Ūˤ,;>ʜļ(.ƐȳŒ-ŌĢ9ʜļ?'
>[27]+A#SPINK5 .dY}Q…F‡̚-ȦȎBɡƈ>),;$'
½Ç+ʝŇB·ƈ'>9-)űA?>[27]<,˟ȓė)ǜʾ>)
SPINK5 kaPDGeyGY(. TEWL ġã>)<[27]HDM ƅð
-ʓȪ1-Ʊ˽.ȓ­,Ŀ> danger signal )ʦʭ?SPINK5 BŸ'
ʓȪ-ɚɹƐȳBɡƈ>),;>ʓȪmƒDǖɺBµˍ>øɺŲ=
Fi,'9üǒ-ǖɺBƶ'>9-)űA?!
ƅðÚǸ 24 Ʈ˪Ŧ,ʽÍˠġãʦ8<?! TIMP1 .ɜɧ|^†t†c
Dˆ\ˮĸĎį̖Tissue inhibitor of metalloproteinase, TIMP̗BTˆf>
˗¨į(=CAD Ƞĥ˙,'ȦȎġŢ?>)ȸ?'>[77]
TIMP1 .ɜɧT‚ˆS‡-ÔʞBƃÙT‚ˆS‡Ǡȳ,;>ɦɡçBµˍ
'á¿ǡȤ,˫>)ȴ<?'>[103]5!AD -źŲȠĥ˙ȅū)?>ɦɡç)ʇȥç, TIMP1 ˫'>øɺŲȸ?'>
[61 117]TIMP rCzƒˆ,. TIMP1 < TIMP4 5(İē?
23
9ǥŲė-ɚɹĦĚʶÔʞ˛ɖ̖Matrix metalloprotease, MMP̗)ʖû­B
ţž>),;$'MMP -˛ɖǥŲBƃÙ>[61, 125]MMP .^‡n
PÔʞ˛ɖ(=MMP1 < MMP14 5( 14 ɀ,Ô̈?'>[61]
TIMP1 .?<?- MMP )9ʖû­Bţžˮĸ>ȅ, MMP9
̖\‚`gˆ\ B̗)-ʛĄŲ̒[103]oe-D…„OˆŲDzȞǒȠĥ˙
,'MMP9 B.8[97]MMP1 ;0 MMP3 - mRNA ʽÍˠ-ġã
ʦ8<?'>):[42]HDM -ÚǸ,;$'R‚`kVFe,>
MMPs mRNA -ʽÍʧŁ?>)B?>)B˥6>)Fi,'
ʥɚ+Ǐʡ.?'+TIMP1 . MMP ,;$'ȓ>ɜɧ¿ĸBƃÙ
'>9-)űA?!
Der f 1 ǭã 24 Ʈ˪Ŧ,'ʽÍˠ-ġãʦ8<?! S100A8 .ɚɹĦ
,İē>L„WG{)ɞûĭ’ȏ;0ìȏ-ˏʸǥŲ)̆ɐȏƚÓɺ
BŒƫ>^‡nPBTˆf>˗¨į(>[22, 121]oe;0FiAD Ƞĥ˙,'.ƼÔį-ȦȎġŢ'>)[114, 126]5!Ƞĥ
˙,.ƒ‡nȏ-¡,ĭ’ȏ;0ìȏ9ħƞʦ8<?>)BɮŻ>)
[47]Der f 1 BŸ!R‚`kVFe<- S100A8 ,;$'ˌɏ>)
µ?'>øɺŲ=HDM ǥŲçR‚`kVFe CAD -ȠŹ,
'˞ʘ+ťàBDŽ!'>9-BűA?!
CAD -ÅȝȠŹ,'.IgE ȔȓB“­)> Th̖Helper T̗2 ė«
-Åȝóů’ŬȨ+ťàBDŽ!'>?<-ÅȝóůBÙŨ>VF
eLF‡;0R}LF‡.“, T ɚɹ<Ȕȓ?>)ɮ<?'!
R‚`kVFe9 T ɚɹ,èƟ>3*ǒ+VFeLF‡:R}LF‡BȔ
ȓ>)oe;0yGY,'ȸ?'>[21, 29]%5=CAD ,
> Th2 ė«-Åȝóů9R‚`kVFe<Ȕȓ?>VFeLF‡
;0R}LF‡,;$'ÙŨ?'>øɺŲ̒Fi,'
24
ʥɚ,Ǐʡ?'+(R‚`kVFe,>Ȕȓěā?'
>VFeLF‡ 31 Ôį,ȳȮDer f 1 ,;$'ȓ>ʽÍĥæ,%'
Ǐʡ!)@IL-8IL-15CCL2CCL20TGF-2 ;0 GMCSF 6 ÔįBTˆf>˗¨į,'ʽÍˠ-ġãʦ8<?!
Der f 1 ǭãŦ,ʽÍˠġã-ʦ8<?! IL-8 .oe- AD Ƞĥ˙,'
ȦȎšˍƬ<)+$'>[53]IL-8 .ĭ’ȏ,Ŀ>ʸçĎį(>3
[1]ĭ’ȏ-ɼ̆ɐ)ʱ̉¯ȕBµˍ>Á>[1, 14]IL-8 - CAD
-ȠŹ,>ťà,%'.ʥɚ,Ǐʡ?'+ĭ’ȏ. CAD Ƞĥ
˙,'9ʦ8<?'>[47]ĭ’ȏ-ˏʸ.mƒDǖɺ-ȶɣ,;$'´
Æ!ɚʊ+*,Ŀ>óů(>)Ƒļ?'>ƼȵɄ-ɞDŽ<
HDM -ÚǸ,;$'9ĭ’ȏ-ˏʸŠʹ?>øɺŲȸ?!
Der f 1 ǭã 24 Ʈ˪Ŧ,ʽÍˠġãʦ8<?! IL-15 .g`€‚„N‚ˆ
̖Natural killer, NK̗ɚɹ;0 T ɚɹ-Ôçġǚ;0ȓİBµˍ>[1]
NK ɚɹ.oe;0Fi- AD Ƞĥ˙,'ʦ8<?>ǻȡŲɚɹ(=
[15, 47]oe(.Ƞĥ˙- NK ɚɹƞ) AD ˞ȡŜ)-Ȱ˫ʦ8<?[19]
ƒȃ+*,;$'ȶɣ!ʓȪ<´Æ!ƅð,Ŀ'óů'>)Ƒļ
?'>[19]‰ƥIL-15 ǓȨ)> T ɚɹ.“, CD8+ T ɚɹ(>
)ȸ?'>[1, 52]Ƽɚɹ9oe;0Fi- AD Ƞĥ˙,'ʦ8<
?[47, 91]IL-15 ,;$'Ƞĥ˙1-ˏʸ)ǥŲçÙŨ?>[63, 106]Ƽ
ȵɄ-ɞDŽ;0?5(-ěāB˥6>)HDM ,ÚǸ?!R‚`kVF
e.ʄǾÅȝɔɚɹ(> NK ɚɹ)ȌŧÅȝɔɚɹ(> T ɚɹBüƮ,ǥ
ŲçȪʀ,>ǻȡBʧŁ'>øɺŲ>)űA?!
Der f 1 ǭã 24 Ʈ˪Ŧ;0 48 Ʈ˪Ŧ)9,ʽÍˠġã-ʦ8<?! CCL2
.ìȏ:yP†rCˆXǔȇɚɹ,'ȦȎ> CCR2 -ƒM‡fBT
ˆf>˗¨į(=[96]oe AD ;0 CAD }d„Ȇ-ȪʀȠĥ˙,
25
'̒ȦȎ'>)ȸ?'>[96, 124]yP†rCˆX.ƅðȅȜŲ
Bƶ+CAD ;0oe AD Ƞĥ˙,'9ħƞʦ8<?>)<
AD ȠŹ,'9®<-ťàBDŽ!'>9-)űA?>[47, 124]yP
†rCˆX.ɚʊ+*-ȜȄBƌƋ>)ǻȡŲVFeLF‡(> TNF-α
: IL-1ĭ’ȏˏʸĎį(> IL-8Th1 ɚɹBǥŲç> IL-12 +*BȔȓ
>)<źŲƸ,> AD Ƞĥ˙(- Th1 óůBµˍ'>9-)ű
A?>[58, 105]‰ƥCAD Ƞĥ˙,'.ǔȇɚɹƞ-ġã-6ěā
?'>[105]oe AD Ƞĥ˙(.ǔȇɚɹƞ-ġã-6+<Thymic
stromal lymphopoietin̖TSLP̗̒ȦȎ'>)9ȸ?'>[86]
TSLP ,;$'ÚǸ?!ǔȇɚɹ.Th2 ėɚɹ-R}LF‡(> CCL17
BȔȓTh2 ėɚɹ<- IL-4IL-5 ;0 IL-13 ȔȓBʧŁ>[86]
!$'HDM ,ÚǸ?!FiR‚`kVFe. CCL2 -ȔȓBŸ'
yP†rCˆX;0ǔȇɚɹBȠĥ˙1ˏʸCAD -ȠŹ,˞ʘ+VF
eLF‡-ȔȓBŠʹ'>øɺŲ>
CCL20 . Der f 1 ǭã 24 Ʈ˪Ŧ;0 48 Ʈ˪Ŧ)9,ʽÍˠġã-ʦ8<
?!R}LF‡BTˆf>˗¨į(=CCR6 -ƒM‡f(>[107]AD
Bþ7D…„OˆŲȪʀȟŴ-Ƞĥ˙,'ġãʦ8<?'> CCR6 Ȧ
Ȏɚɹ.ƺžȁ+ǔȇɚɹ;0‚‡S„l‡Yɚɹ(>)ȸ?'
>[107]CCL20 .oe AD Ƞĥ˙(ȦȎšˍ'>3ʑǮ’-ǺŜ)˞
ȡŜ)Ȱ˫>)ȸ?'>[82]Deqˆna`cYe,'˳Ų
óůʦ8<?!oe-Ȫʀ(. CCL20 mRNA -ʽÍˠġã))9,
CCR6+‚‡S„l‡Yɚɹ-ġãʦ8<?'>[38]oe;0Fi- AD
Ƞĥ˙,'.ġã!‚‡S„l‡Yɚɹ, IgE ɞû'>)
ȸ?'>)<‚‡S„l‡Yɚɹ.ƅðȅȜȨ+ IgE BŸ'ȅȜ
ȨƅðBƌƋ>)ɮ<?'>[9, 90]ƅð,;$'ǥŲç!‚‡S„l
26
‡Yɚɹ. IL-1βIL-6IL-12TNF-α ;0F‡^ˆrH†‡BȔȓ>
)oe;0yGY,'ȸ?'>)<[5]̢ė-6+<ė
D…„Oˆóů,9˞ʘ+ťàBDŽ!'>)űA?>žč-ɞDŽ)?
5(-ěā<HDM ǥŲçR‚`kVFe.CCL20 BȔȓ>),;
$'‚‡S„l‡Yɚɹ-Ƞĥ˙1-ˏʸBµˍVFeLF‡-Ȕȓ
BŸ'ȌŧÅȝ1-ȾʒBµˍ'>)ɮ<?!
Der f 1 ǭã 48 Ʈ˪Ŧ,ʽÍˠ-ġãʦ8<?! TGF-2 .ƃÙŲVFe
LF‡(=-3,9 4 %-DF]rJˆ{İē>)oe,
'ȸ?'>[36]TGF-β2 . TGF-β1 )ɕ 70̕-ȰüŲBƶǖɺ9
̈ª>)oe;0yGY,'ȸ?'>[36]<,oe,
'.T ɚɹ- IL-4 ;0 IL-5 -Ȕȓ) B ɚɹ-ġǚBƃÙ>)9Ƭ
<,+$'>[30, 62]oe;0Fi- AD ȠŹ,> TGF-β2 -ťà
,%'.Ƭ<,+$'+CAD Ƞĥ˙,> TGF-β -ȦȎ¬
ȸ?'>)<[87]T ɚɹǖɺ-ƃÙǖǑ-ȶɣ AD ȠŹ,˫
'>øɺŲ.>)űA?>[73]žč-ȵɄɞDŽ<HDM ,;$'ǥŲ
ç?!R‚`kVFe.TGF-β2 BȔȓ>),;=T ;0 B ɚɹǖɺBƃÙȌŧÅȝɔ-ǻȡóůBƃÙ'>øɺŲȸ?!
Der f 1 ǭãŦ,ʽÍˠġãʦ8<?! GM-CSF .ħɺŲˊʑŗɚɹB
ĭ’ȏìȏ5!.ĭ˜ȏ,?
?Ôç>ǖɺBƈ%VFeLF‡BT
ˆf>[1]GM-CSF .oe AD Ƞĥ˙(-ȦȎġãʦ8<?'=[12]
Ƞĥ˙(ġã-ʦ8<?'>ĭ˜ȏ-Ôç;0ǥŲç,;>ɼ̆ɐ,˫
>3[59]źŲ-Ƞĥ˙,>yP†rCˆX-DxeˆWYˮĸBŸ
'źŲǻȡBɡƈ'>)ȸ?'>[12]5!GM-CSF .oe
AD Ƞĥ˙,>R‚`kVFe<-Ȕȓġãȸ?'=Ƞĥ˙,
>‚‡S„l‡Yɚɹ;0ǔȇɚɹ-ġã;0ǥŲç1-˫ȸć
27
?'>[92]CAD Ƞĥ˙,> GM-CSF -ťà,˫>ʥɚ+Ǐʡ.ʒA
?'+GM-CSF -ȪƄ.ȕˠ²İȨ,ĭ’ȏìȏ;0ĭ˜
ȏ-ġãBʧŁ>)<[83]CAD Ƞĥ˙,>ǻȡɚɹ-ġã,ƼÔ
į˫'>)ɮ<?>[47]<,Fi-ƻǍʑìȏB GM-CSF (
ÚǸ>)“ʘɜɧ˕ûƅðʖû­̖Major histocompatibility complexMHC̗
class II BȦȎ>ǔȇɚɹġã>)<[115]ƼÔį.ǔȇɚɹ1Ôçžȁ;0ǥŲçBŸ'CAD Ƞĥ˙,>‚‡S„l‡Yɚɹġã,˫'>)ɮ<?'>[91]žč-ȵɄ-ɞDŽ<HDM ,;
$'ÚǸ?!R‚`kVFe.GM-CSF BȔȓ>),;$'ĭ’ȏ
ìȏ;0ĭ˜ȏ-ġãBʧŁ>))9,‚‡S„l‡Yɚɹ:ǔȇɚɹ
-ǥŲçBˉ'ȌŧÅȝ1-ȾʒBµˍ'>)ɮ<?!
CAD -ȠŹ,'.ƅðȅȜȨ IgE B’Ŭ)> I ėːƛȡ˞ʘ+ť
àBDŽ!t‚Zyɚɹ,>ƅðȅȜȨ IgE -Ȕȓ. Th2 ɚɹ<-V
FeLF‡ŭ̅(=- Th2 ɚɹ-Ôç;0ǥŲç,.‚‡S„l‡
Yɚɹ,;>ƅðƓȸŭʘ)+>!$'CAD -ȦȡǖǑ-ǐ8'×
Ƹ-Ǜ˴,'.‚‡S„l‡YɚɹB’Ŭ)!ȌŧÅȝɔ-Ȝœ+ǥ
Ųçȓ'>øɺŲ>‚‡S„l‡Yɚɹ.ƺžȁ+ȇŹ(ʓȪË
,İē>ƅðƌƋÔç>)ƒ‡nɍ1ˏʸgFˆs T ɚɹ,ƅ
ðBƓȸ>[66]ƼȵɄ,' mRNA ʽÍˠġãʦ8<?! GM-CSF
.‚‡S„l‡Yɚɹ-žȁǥŲç;0Û̏ɚɹ<-Ôç,˞ʘ+V
FeLF‡(>)B˥6>)[20]HDM ǥŲçR‚`kVFe<Ȕȓ
?!ħˠ- GM-CSF ǐ8'×Ƹ- AD ÅȝȠŹ,˫'>)ɮ<?!
!$'HDM ǥŲçFiR‚`kVFe,> GM-CSF -ȔȓǖǑ
Ƭ<,+?/CAD ,Ŀ>ƤʚǡȣǢB˩Ȧ(>øɺŲ>)ɮ!
(ǘ-ɉ—ɇ(.FiR‚`kVFe,> GM-CSF ^‡nP-Ȕȓ
28
,%'ʣ³>))!
29
1 .5. ʓ
30
kŠªÙÒµ’•‡J{´­¬±LG®m·º¼±¬»z
z
AQP3
Carbonic anhydrase II
CCL13/MCP4
CCL17/TARC
CCL18/PARC
CCL27/CTACK
CD1a
CD26 dipeptidylpeptidase IV
CD45
Chondroitin sulphate proteoglican 2
Collagen VI a-1 C-terminal globular domain
CXCL8/IL-8
CXCL10
Cysteine-rich protein 1
Cysteine-rich protein 2
Elafin
Fibronectin 1
Filaggrin
IL-1B
IL-1F9
Involucrin
Keratin 1
Keratin 2A
Keratin 5
Keratin 6A
Keratin 6B
Keratin 10
Keratin 14
Keratin 16
Keratin 17
Loricrin
Lymphocyte-specific protein 1
N-myc downstream regulated gene 1
Nel-Like 2
Osteoblast specific factor 2
Plasminogen activator inhibitor, type I
PPP1R5
S100A2
S100A7
S100A8
S100A9
Serine protease inhibitor clade B, member 13
Small proline-rich protein 1A
Small proline-rich protein 1B
Small proline-rich protein 2B
Small proline-rich protein 2C
Small proline-rich protein 3
Tenascin precursor
Trichohyalin
VNN1
VNN3
β-defensin 2
:c
>ÐàÕã
B}d>|X
MiHyq
‡Zb­¹¶Hµyq
âåØH­¹¶9EZbµyq
ONZbµyq
9EZb­¹¶‡“‡ZbµfhM‡
‡Zbkƒ-éÍâåÛäÑ¿æÎ
MiHv-
Zbp«Q\ZbrYµ~:c
ž—‡ÉáæÈå½ÉæÓ°»‡”‡3SHEÓÞÀå
Hyq
ÂÚÁÅÏ懝‡Zbyq
ZbpéZbËÆÔã
ZbpéZbËÆÔã
ÂáÌÏæ΁!é-(ID?)
Zb.P«Zbµ*~«yq«
ÇáÐÖÊÀÒµ«ÇáÐå_^µ‚
AK)ÊÀÒÃÀå«âåØH?)
—˜Š‡Ú¾ÝâæµêR
l#­¹¶l#t$µ7whM
16#ÇáÐÖÊÀÒµZb…5痗ÇáÐåè
NZb´LG°»‡——‡ÇáÐå
&#ÇáÐÖÊÀÒµZb…5痗ÇáÐåè
+%NZbµZb…5痗ÇáÐåè
+%NZbµZb…5痗ÇáÐåè
16#ÇáÐÖÊÀÒµZb…5症ÇáÐåè
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l#­¹¶C´LG°»‡—‡ÇáÐå
#eNZb´LG°»‡—‡ÇáÐå
l#­¹¶l#t$µ7whM
–ŠŸ ¨¢£‡³]«Zb…5½'*
Zbp³4µµËÆÔã²<
;³Zb½oU
N*~
NZbµ.P³yq雡¦¢¤§¨¢£‹›šœ™
ÛáÌÝÖæÈå?)! «_@½,
hMdâå}|X
Zb2xj«Zb
”•ˆ‡âåØH­¹¶Hµq) ‹¥§¤¦¢Ÿ§¢£
Hµ?)«¿áÄÓå}su
œŒŒ’ë³³¸´„WMiH­¹¶H´LG
ÄßÒâÛËå8ÍâåÛäÑ¿æ΁!
ç0è
ç0è
ç0è
l#t$µ7**«×â¿:c´€ì
ç0è
Zb.P«<T´€¯«[`7V½x
=´LG«=†µ<
™–Š©‰‡—˜Š‰‡—˜‰‡—˜Š‘‡­¹¶‡—˜ŠŽŽ‡½n"
™–Š©‰‡—˜Š‰‡—˜‰‡—˜Š‘‡­¹¶‡—˜ŠŽŽ‡½n"
-gÜÛÐÓ
31
a/F
[82]
[52, 82]
[82]
[52]
[82]
[82]
[82]
[82]
[52]
[109]
[82]
[52]
[52]
[82]
[82]
[52, 109]
[109]
[109]
[52]
[109]
[109]
[109]
[109]
[109]
[109]
[109]
[109]
[109]
[109]
[109]
[109]
[60]
[60]
[52, 109]
[82]
[82]
[82]
[109]
[109]
[109]
[109]
[82]
[109]
[109]
[109]
[109]
[109]
[82]
[109]
[52]
[52]
[52, 109]
Ja_buecf^5W€yx}72zO‚…ˆ}x‡V
V
+F
B$1
ARTS-1
" Ÿ˜›ƒ‘‹š‹©‰%
[74]
POSTN
9?E#:UQ`jmqnprsno`jiklh
[74, 121]
Down-regulated in ovarian cancer 1 isoform 2
«&¬
[74]
eIF-5B
ANZL
[74]
Kinectin 1
£ª—ªI8P~w‡œ’©
[74]
Myosin Va
£ª—ªI8P
[74]
A-kinase anchor protein 9 isoform 2
«&¬
[74]
Canis familiaris ret proto-oncogene (RET)
«&¬
[74]
GOLGA4 subfamily a, 5
”¥¡›RS
[74]
GOLGA4 subfamily a, 4
”¥¡›RS
[74]
GOLGA4 subfamily a, 5
”¥¡›RS
[74]
EEA1
(Œ©›–ª¢L=@{,‚
[74]
CG1574-PA
«&¬
[74]
chromosome 6 open reading frame 142
«&¬
[74]
sushi-repeat-
«&¬
[74]
Hypothetical protein LOC55086
«&¬
[74]
S100A8
3-vŠ¥›©XRS
[74, 121]
INPPL1
d^?E­C/?Ev¥©§©“?E-
[74, 121]
SCCA-2
£š¦Ÿ’©*”¦©Ÿ¨™Šª•\
[74]
SAA3
I8
[74]
Serum amyloid A protin (Canis familiaris)
I8
[74]
SAA
I8
[74, 121]
TIMP1
?EPM‰!
[74]
similar to RAD50 homologue isoform 1
«&¬
[74]
CIDE-3
GD?E
[74]
sperm-associated antigen 5 (Astrin)
)H'<K>Y[TI8P
[74]
Mucin-15
;. [74]
hypoxia-induced gene 1
«&¬
[74]
DPP4/CD26
l^?EJ]"®”¦©Ÿ¨™Šª•
[121]
SPINK5
”¦©Ÿ¨™Šª•\
[121]
SGPL1
?EUQƒ?E€[|‡^kaj^‰
%€„† [121]
PPAR
hg_td^-!€„‡"064
[121]
Cullin4A
¤ž˜©¦Žª•
[121]
32
1-3
TNF-α
IFN-γ
IL-1
IL-12
IL-6
IL-8/CXCL8
IL-7
IL-10
IL-15
IL-18
TGF-β
CCL2/MIP-1
CCL5/RANTES
CCL7/MCP-3
CCL8/MCP-2
CCL11/Eotaxin
CCL12/MCP-5
CCL13/MCP-4
CCL17/TARC
CCL20/MIP-3α
CCL22/MDC
CCL27/CTACK
CXCL9/Mig
CXCL10/IP10
CXCL11/IP9
α-MSH
PGE2
GM-CSF
SCF
7
"+jeohewX/36_'\ X36\dsoxkm9
tivqcxlY`]$36\ "+jeohewX/36\ UNEB36^\9XSPB36Y`]BUB36[ZaBOMSDW9
)(36\<
'=;
KB36\A36\<
tivqcxlY`]$36\BOQDEFB)(XRNLBOOB-&<
SPB36Y`]BUB36\<
SPB36Y`]BUB36\BOMSDW9
UB36\Y`]7\y:36\BOVJB)(
LLTFBb-&Za
'\=;
LLTECBLLTGCBLLTIBb-&Za.8'\=;
LLTECBLLTFCBLLTGBb-&Za
'X>'Y`]'\=;
LLTECBLLTFBb-&Za5!36X>'X'X
'XU36\=;
LLTGBb-&Za>'X'yU36\=;
LLTFBb-&Za
'X>'\=;
LLTFCBLLTGBb-&Za
'X>'X'XU36\=;
LLTHBb-&ZaBUNFB36Y`]'\=;
'236\=;
LLTHBb-&Za
'_rupxU36\=;
0/36\=;
gqfinx36\=;
gqfinx36\=;
gqfinx36\=;
"+*
"+X-#X-,
'X@1'Y`]$36\X Y`]
36?
33
4%
[1,21,29,37]
[1,29]
[1,21,29,37]
[1,21,29,37]
[1,21,29,37]
[1,21,26]
[21,29,37]
[1,21,29,37]
[1,21,29,37]
[1,21,29,37]
[1,21,29,37]
[29]
[1,21,37]
[29]
[29]
[29]
[29]
[29]
[1,21,29]
[1,21,29]
[29]
[1,21,29]
[1,21,29]
[1,21,29]
[1,21,29]
[21]
[1,21,29]
[21,29]
[1,21,29]
34
1-5
26
58
603
687
65
179
1566
1810
623
29
5
1659
247
32
657
1938
1344
3748
35
36
523:192
Aquaporin 3
718426()'$(
<
./
!
!
2.263 **
14.152 **
2.168
3.382 **
1.296
NA
NA
1.257
0.127 *
0.474 *
0.132
0.672
NA
NA
4.882 **
1.055
1.406
1.066
NA
NA
11.486 **
3.084 **
19.973
12.149 **
8.562 **
11.860 **
1.102
NA
NA
0.964
NA
NA
0.604
0.817
0.800
NA
NA
0.375 *
0.288 *
0.373 *
4.198 **
NA
NA
0.771
NA
NA
0.627
NA
NA
1.411
NA
NA
1.321
0.629
NA
NA
0.567
0.318 *
0.831
0.279
0.462 *
0.468 *
2.223 **
0.559
0.766
NA
NA
0.771
1.065
NA
NA
ND
NA
NA
";.
&%"+0,**&#"-,
8.725 **
9.657 **
PREDICTED CA2
XM_535117
0.530
Carbonic anhydrase II
0.543
NM_001003051
0.547
CCL17
CCL17
NM_001003968
1.144
CCL27
CCL27
PREDICTED DPP4
XM_535933
ND
Dipeptidyl peptidase 4 / CD26
1.076
partial Collagen VI-alpha1
COL6A1 AJ388539
NM_001003200
2.143 **
IL-8
IL8
2.389 **
2.185 **
2.123 **
DR104583
0.988
Cysteine-rich protein 1
CRIP1
U52105
0.836
Fibronectin (Fragment)
U16207
0.831
XM_536059
0.859
PREDICTED KRT5
XM_855013
1.157
Keratin, type II cytoskeletal 5
1.101
1.007
1.063
NM_001013425
1.666
keratin 10
KRT10
PREDICTED KRT16
XM_548101
0.919
Keratin, type I cytoskeletal 16
XM_849236
0.795
lymphocyte-specific protein 1 isoform 1 PREDICTED LSP1
PREDICTED NDRG1 XM_539170
1.421
N-myc downstream regulated gene 1
1.388
PREDICTED SERBP1 XM_536673
0.989
Plasminogen activator inhibitor 1
RNA-binding protein
0.657
PREDICTED S100A2
XM_850065
0.751
S100 calcium-binding protein A2
1.104
1.113
S100 calcium binding protein A8
S100A8 DR104918
PREDICTED S100A9
XM_547585
1.102
S100 calcium-binding protein A9
1.123
XM_538811
ND
Tenascin precursor, transcript variant 1 PREDICTED TNC
NM_001003372
0.757
vanin 1
VNN1
".
Genbank
Accession
Number
PREDICTED AQP3
XM_849503
1-6
AD *
**
*
**
37
PREDICTED SGPL1
Sphingosine-1-phosphate lyase 1
Peroxisome proliferator-activated receptor gamma
PPARG
cullin 4A isoform 1
PREDICTED CUL4A
DPP4
SPINK5
Dipeptidyl peptidase 4 / CD26
Serine peptidase inhibitor, Kazal type 5
PREDICTED SRPX2
PREDICTED INPPL1
sushi-repeat-containing protein
Inositol polyphosphate phosphatase-like 1
SAA1
S100A8
TIMP1
PREDICTED MYO5A
PREDICTED AKAP9
PREDICTED EEA1
Myosin-5A (Myosin Va)
A-kinase anchor protein 9 isoform 2
Early endosome antigen 1
Serum amyloid A protein
S100 calcium binding protein A8
TIMP metallopeptidase inhibitor 1
PREDICTED KTN1
Kinectin 1
PREDICTED FILIP1L
Downregulated in ovarian cancer 1 isoform 2
**
**
**
**
**
**
**
**
**
**
**
**
**
*
**
**
*
2.075 **
NA
NA
1.055
0.166 *
1.08
NA
NA
1.406
172.01 **
ND
0.559
1.162
NA
NA
NA
NA
ND
2.223 **
3.178 **
NA
NA
NA
NA
NA
NA
NA
NA
!
NA
NA
<
./
XM_535715
&%"+0,**&#"-,
!
15.024
15.136
15.542
1.766
1.621
1.655
1.209
0.834
0.467
0.717
1.203
1.019
0.512
2.177
1.216
2.446
3.651
2.398
5.664
0.468
0.765
0.841
4.882
2.220
2.448
2.390
2.269
3.407
0.755
1.083
1.023
1.044
718426()'$(
";.
**
**
**
523:192
2.195
2.277
2.640
1.035
XM_537455
1.493
1.141
1.165
0.870
XM_535487
1.883
XM_532456
0.876
XM_532649
1.113
1.030
0.633
XM_549134
1.782
XM_542327
1.155
2.672
0.537
1.609
3.679
NM_001003050
1.113
DR104918
1.379
NM_001003182
1.236
ND
XM_535933
1.992
NM_001025397
1.719
3.890
2.017
1.323
XM_546150
0.735
NM_001024632
0.880
XM_844449
0.916
0.977
".
Genbank
Accession
Number
1-7CAD 38
TGFB2
CSF2
Transforming growth factor beta 2 (Fragment)
Granulocyte-macrophage colony stimulating factor
IL15
IL15
IL18
Interleukin-15 (Fragment)
Interleukin 15
Interleukin 18
CCL2
CCL17
CCL20
CCL27
TGFB1
IL7
Interleukin 7
CCL2
CCL17
CCL20
CCL27
Transforming growth factor, beta 1
IL12A
IL12B
IL8
Interleukin 12A (IL-12, p35)
Interleukin 12B (IL-12, p40)
Interleukin 8
PREDICTED
TNF
Tumor necrosis factor
523:192
!
0.823
1.890
1.775
ND
11.486
12.149
8.562
11.860
ND
0.870
2.025
1.751
0.622
0.601
4.388
0.127
19.046
0.672
1.020
1.028
2.050
2.170
6.806
4.213
&%"+0,**&#"-,
NM_001003244
ND
ND
NM_001003293
1.747
NM_001003292
ND
NM_001003200
2.143 **
2.389 **
2.185 **
2.123 **
NM_001048138
ND
1.144
AF479882
1.985
XM_844053
1.642
NM_001003169
0.835
0.840
NM_001003297
3.755 **
NM_001003051
0.547
NM_001005254
ND
NM_001003968
1.144
NM_001003309
1.041
1.043
XM_853503
1.180
DQ525400
1.530
NM_001003245
2.474 **
ND
".
Genbank
Accession
Number
6.376 **
0.474 *
9.530 **
NA
NA
2.199 **
NA
28.470 **
0.132 *
12.424 **
NA
NA
1.096
NA
NA
NA
<
./
**
1.806
3.778 **
**
**
2.278 **
9.996 **
**
";.
**
*
**
**
**
**
**
**
NA
NA
19.973 **
!
NA
NA
NA
3.084 **
NA
718426()'$(
1-8
ɉ ̛ɇ
"%!$&
# GM-CSF 39
2 .1. Śʫ
GM-CSF.̆ɐȏìȏǔȇɚɹ;0‚‡S„l‡Yɚɹ+*-Ôç
;0žȁBµˍ>VFeLF‡)'Ȧʙ?![16]Ȏē(.oe,
'ĭ˜ȏ-ǥŲçBµˍ>)9Ƭ<)+$'=[116]ǻȡŲVFeL
F‡)'ǒ+ȟŴ-ȠŹ,˫'>9-)ɮ<?'>
ƼÔį.oe-˫ɍƒGy`ǻȡŲɾȟŴĊų;0źŲ˨ğŲɶȟŴ
+*ʖƞ-ǻȡ-ɖĎ,˫'>)ɮ<?'>[76]˫ɍǷʁǻB“
Ƞĥ)>ʄŏÅȝŲȟŴ(>oe-ƒGy`,'.ǷʁËʑɌĂĐ,
ƅðƓȸɺBƶ>ǔȇɚɹÓȎ̡ɚɹ;0̣ɚɹ-ǥŲçȓ'
> [24] ƒ G y ` , Ŀ > ƅ GM-CSF ö ĺ ­ ƅ ­ ( > Mavrilimumab
̖Medimmune Gaithersburg MD̗-Ƅ,;$'ʃřȡȇƙĉ>
)<[17]GM-CSFƼȟŴ-ȠŹ,ǫ˫'>9-)űA?>D…
„OˆŲȟŴ(.ȑĠƅð,Ŀ>IgEƅ­-Ȕȓ,˫A>ǔȇɚɹ̣ɚɹ
;0̡ɚɹ˫'>Ǽ(ƒGy`-ȠŹ,̈ª'>ǔȇɚɹ-Ôç
žȁ;0ǥŲç,;>ƅðƓȸɺ-ʵǥ,GM-CSF˞ʘ(>)<
[120]?<-ȟŴ,'ġã>GM-CSF.ƅ­Ȕȓ,!>ǐ8'×ƸȠŹ,˫'>øɺŲ>GM-CSF-“ʘ+Ȕȓɚɹ)'̣ɚɹ
;0yP†rCˆXİē>oeADŴɯ<Ǝõ!ʓȪR‚`kVFe
-×£ę̍,'.½œɯȗǁ-R‚`kVFe,ǜ2'PMA,;$'ʧŁ
?>GM-CSFȔȓˠƶŷ,ġã!)ȸ?'>)<[92]R‚
`kVFeȔȓ>GM-CSFAD-ȠŹ,˫'>øɺŲ.̒
Der f;0Der p.oe;0Fi-AD,>“ʘ+ȑĠD…„S‡(
>ȅ,Der f.Fi-AD,'Ƶ9ħŸ¯ʦ8<?>HDM(>
)ȸ?'>[46]Der f-“ʘ+D…„S‡^‡nP(>Der f 1.nnF
40
‡ǒWYcF‡t†cDˆ\(=[28]ƼÔį-ƶ>^‡nPʶÔʞǥŲ.
HDM-D…„S‡Ų)˫ˋ>)ȸć?'>[75]oe-ǙœȪʀ-R
‚`kVFe,'.HDMD…„S‡ÚǸ,;$'GM-CSFȔȓµˍ?
-Ȕȓ.t†cDˆ\ˮĸÞ,;$'ƃÙ?!)<[88]HDMD…„S
‡-^‡nPÔʞǥŲGM-CSFȔȓ,˞ʘ(>)ɮ<?'>ɉ1ɇ-ɞ
DŽ;=Ȇ,'9HDMD…„S‡,;$'R‚`kVFe,>GM-CSF
mRNA-ʽÍʧŁ?!)<R‚`kVFe<Ȕȓ?!GM-CSF
Ȇ-D…„OˆŲǻȡBʧŁ'>øɺŲɮ<?!+<
GM-CSF-^‡nP…v„,>ȦȎ,%'.Ǐʡ?'+!8Ƽɇ
(.Der f-ǭã,;$'R‚`kVFe<GM-CSF^‡nPȔȓ?>
,%'Ǐʡ!
41
2 .2. ƾƢ)ƥǢ
2.2.1. ƾƢ
F i ʓ Ȫ R ‚ ` k V F e - ɚ ɹ NJ ( > CPEK ) - ę ̍ Ǫ CnT-09 . CellnTEC advanced cell system<ʷÆ!5!ǽʑǮęĕOpti-MEM I
̖Invitrogen̗;0ɠ£ę̍Ʈ-ɚɹÝ˺,TrypLE Express̖Invitrogen̗B
° ȕ ! DPBS ; 0 W Y c F ‡ t † c D ˆ \ ˮ ĸ Þ ( >
Trans-Epoxysuccinyl- L-Leucylamido- ̖ 4-Guanidino ̗ Butane ̖ E-64 ̗ .
Sigma-Aldrich̖St. Louis, MO̗<ÆƂ!HDM)'-Der f-ƆÓȄ.
Greer Laboratories̖Lenoir, NC̗<ʰģ?'>9-Bȕ!
2.2.2. ɚɹę̍
̞č<10čɠ£!CPEKB10 mL-CnT-09,Ǩˏ375% CO2ȑĠ(75 cm2r‚YT̖NUNC̗Bȕ'ę̍!80%T‡r„I‡e,˒
!Ǜ˴(ę̍ǪB˰ñ10 mL-DPBS(ɚɹBǤǧ!TrypLE Express
B3 mLã!Ŧ5-10Ô˪ 37,˾ɨ'ɚɹBÝ˺!CnT-09B7 mL
ãɕ80̕-ɚɹÝ˺Ǩˏ!)Bȷʦ!ŦɚɹǨˏǪB15 mL
centrifuge tube̖NUNC̗,čôķǰ( 180 G5Ô˪˔Ŭ!˔ŬŦ
ŒǮB˰ñɚɹĝBCnT-09Bȕ'ÌŽǹ!ÌŽǹ!ɚɹǨˏǪ
(2×104/mL)B1 mL%24-well plates̖NUNC̗,Ôǣ80̕T‡r„I‡e
,˒>5( 37 5%CO2-ǀ¥(ę̍!-Ŧę̍ǪBOpti-MEM I
,ĥƳǽʑǮ(24Ʈ˪ę̍Der f-ǭãĶ̐Bʒ$!
ǭã!Der f.“ʘD…„S‡(>Der f 1-þƶˠƬȷ+•ȂÏɞą(
Der f 1)'10 μg/dL)+>;DPBS(ǵʞ!Der f 1ǵʞǪ̖0.1, 1;
42
010 μg/mL̗5!.DPBSB10 μL%úGH„,ǭã245!.48Ʈ˪ę̍
!
t†cDˆ\ˮĸÞ-ǭãĶ̐(.DPBS(ǵʞ!E-64ǵʞǪBDer f 1ǵ
ʞǪ,ã'37(15Ô˪ÑȐ!CPEK<ǽʑǮę̍ǪB˰ñDer f
1̖10 μg/mL̗;0E-64̖10-7 ̤ 10-9 M̗Bǭã!ę̍Ǫ,ĥƳ37
5%CO (48Ʈ˪ę̍!
2
2 .2.3. ɚɹȓİȍ
t†cDˆ\ˮĸÞ;0Der f-ǭãɚɹ-ţŹ;0ȓİȍ,>Ť
́Bʣ³>!8,CPEK,?
?Bǭã'48Ʈ˪ę̍!ŦɚɹţŹ
B»Ɇ̇Ūˤ(ʜļ>))9,Trypan blue 0.4%̖Sigma-Aldrich̗Bȕ
'ɚɹȓİȍBǏʡ!
2.2.4. ę̍ŒǮ,>GM-CSFǺŜ
CPEKBDer f5!.E-64;0Der f-İē(24Ʈ˪5!.48Ʈ˪ę̍!
Ŧę̍ŒǮBčô!"#,-80(Ïɞ·İ!ę̍ŒǮ’-GM-CSFǺ
Ŝ. DuoSet® Canine GM-CSF ELISA kit̖R&D System, Minneapolis, MN̗
Bȕ'DZĵ!ǓǴƲɦ.Der f5!.E-64;0Der fBã!ę̍ŒǮ
,Nae,¢ņ-ƒT‡pg‡eFiGM-CSFBő˝'¯ž!
2.2.5. ɟʠ
ȈɆ!Ķ̐B3čĶƦę̍ŒǮ,>GM-CSFǺŜ-ŕĖ¼BɋÓ!
úDer fǺŜ,>GM-CSFȔȓˠ-ƶŷŎBʣ³>!8,Tukey-KramerHSDǏĵBȕ!t†cDˆ\ˮĸĶ̐(.E-64Bǭã'+Der fǭ
ãɬBĿǿ)!Dunnet ǏĵBȕ!ɟʠIJȨƶŷŎ. P < 0.01 )ĵɭ
43
!ʞǃ,.ɟʠ]re JMP version 5.1.2̖SAS institute, Cary, NC̗B°
ȕ!
44
2 .3. ɞDŽ
2.3.1. CPEK-ɚɹţŹ)ȓİȍ
Der f;0E-64-İē,'9ɚɹ-ţŹ.ǽǭãɬ)Ȱ˓+ɗˢţ
Bȸɚɹ˪;0W~ˆ…ś˙1-ƐȳŲ9ɡƈ?'!̖đ2-1̗5!
?-ǀ¥,'9ɚɹȓİȍ. 79.7̤84.6% (=Der f;0E-64
ǭã-Ť́.ʦ8<?+$!̖đ2-2̗
2.3.2. Der fǭã,;>R‚`kVFe-GM-CSFȔȓ
ę̍24Ʈ˪Ŧ,>ę̍ŒǮ’-GM-CSFǺŜ.ǽǭãɬ 2.042 ±2.095
pg/mL Der f 1ǭã 1 μg/mLɬ 5.273 ±6.371 pg/mL Der f 1ǭã 10 μg/mL
ɬ(. 21.733 ±18.551 pg/mL ($!ę̍48Ʈ˪Ŧ,'úɬ-ę̍ŒǮ
’-GM-CSFǺŜ.ġãÀýBȸǽǭãɬ 5.214 ±3.985 pg/mLDer f 1
ǭã 1μg/mLɬ(. 40.156 ±11.131 pg/mL10μg/mLɬ(. 70.415 ±19.169
pg/mL($!ɟʠʞǃ-ɞDŽCPEK-GM-CSFȔȓ.Der f 1-ǺŜ;
0ę̍Ʈ˪,²İ'ƶŷ,ġã>)Ƭ<)+$!̖đ2-3̗
2.3.3. WYcF‡t†cDˆ\,;>GM-CSFȔȓ-ƃÙ
Der f 1BŸ!R‚`kVFe-GM-CSFȔȓ.E-64B10-7 M-ǺŜ(ǭã
!Ĝû(-6ƃÙ?10-8 M¤-ǺŜ(.Ŀǿɬ̖Der f 1-6̗)-ƶŷ
Ŏ.ʦ8<?+$!̖đ2-4̗
45
2 .4. ɮļ
ƼȵɄ-ɞDŽ<Der f .Fi-R‚`kVFe,Ŀ'9 GM-CSF -Ȕ
ȓBʧŁ>)Ƭ<)+$!5!GM-CSF -Ȕȓ.WYcF‡t†
cDˆ\ˮĸÞ,;$'ƃÙ?!)<ȔȓǖǑ,. Der f -t†cDˆ
\ǥŲ˫'>)űA?!Der f ,.WYcF‡t†cDˆ\(>
Der f 1 þ5?'=CPEK ,> GM-CSF -Ȕȓ,'“ʘ+ťà
BDŽ!'>)ɮ<?!HDM -ƅð^‡nP(> Der f 1 .ńËĘ 1 g ’,'. ŕĖ 2.25 ±4.97 μg İē'=ħ)@(. 30 μg
¤Œİē>Ĝû9>[99]5!CAD ɩŴȆB̌ɵ>ĹńË-Ę,İē
> Der f .̌ɵȆ CAD ,ɩŴ'+Ĺŝ)<2'ħ)ȸ
?'>)<[31]ƼȵɄ(ȕ! Der f 1 ǺŜ̖10 μg/mL̗.Ķ˶-ȑ
Ġ’)34üȿŜ-ǺŜ(>)ɮ<?!
HDM ȅȜȨ IgE -Ȕȓ,ÃɆ#ƅðƓȸɚɹ,;>ƅðƌƋ)ƒ‡nȏ
1-ƅðƓȸŭʘ(>ʓȪ,>ƅðƓȸɚɹ(>‚‡S„l‡Y
ɚɹ.ƺÔçȇŹ(ʓȪ,İēÚǸ,;$'ǥŲçMHC class II B
ȦȎ'ƅðBƓȸ>[69]ǥŲç!‚‡S„l‡Yɚɹ.ʓȪ-^Fe
X~‡PW‡BʦʭÔDžBʝŇƥý,©˦>),;$'ƅðBƌƋ
>)yGY,'Ƭ<,?'>[69]-;, GM-CSF .‚
‡S„l‡Yɚɹ-Ôç;0ƅðƌƋɺ-ǥŲç,˞ʘ+Ôį(>
GM-CSF -“ʘȔȓɚɹ. T ɚɹ:yP†rCˆX+*(>ƼȵɄ-ɞ
DŽ<ƅð´Æ-ǐ8'ƪƸ-Ǜ˴,'˷Ɛ>R‚`kVFe<Ȕ
ȓ?>øɺŲ̒5$!oe,'.PMA̖Phorbol 12-myristate
13-acetate̗ÚǸ,Ŀ>ʓȪR‚`kVFe- GM-CSF Ȕȓ-ˠ AD Ŵɯ
)½œɯ,'Ȝ+>) in vitro (ȸ?'>[92]!$'
46
GM-CSF Ȕȓ|LhZ{BƬ<,>)üƮ,½œȆ) CAD ɩŴȆ-R
‚`kVFe,> GM-CSF Ȕȓˠ5!.Ȕȓ|LhZ{-ǜʾǏʡ+<
0,˗¨įĥȜ-ƶǽ,%'ǏʡCAD -ȠŹʞƬ-6+<R‚`kV
FeBǓȨ)! CAD ,Ŀ>ƤʚǡȣǢ-˩Ȧ,ƶȕ(>)űA?!
WYcF‡t†cDˆ\BŸ! CPEK -ǥŲçǖŚ)'Protease
activated receptor̖PAR̗-˫ɮ<?>PAR .ȅĵ-t†cDˆ\,
;$'ȅȜȨ,ǥŲç?>‹ˠ­ G ^‡nPÉť-̟čʁʲˉėöĺ­(
>Ȏē5(,̝%- PAR rCzƒˆüĵ?'=-rCzƒˆ-‰
%(> PAR-2 .ȓ­Ë,ŘÔŐʓȪĚśŇ-R‚`kVFe,'
9ȦȎ'>)ȸ?'>[49]PAR-2 -ɚɹʁĦ,˽Ó'> N
ƻɈ¾wt`fˣ.eƒtW‡eƒt^ˆ\5!.ʑǪÐĒɉ VIIaXa Ďį
Ɋ-t†cDˆ\,;$'Õƣ?>Õƣ?! N ƻɈ. Tethered ligand
)' PAR-2 Ôį-ɚɹĦɉ̛„ˆt,ɞûWQg„¨˒?>PAR-2
.“,[ƒ‡t†cDˆ\,;$'ǥŲç?>)ɮ<?'!˄Ŗ
oe-ɚɹNJ,'WYcF‡t†cDˆ\,;$'9ǥŲç?>)
Ƭ<)+$!)<[100]Der f 1 ,;> CPEK -ǥŲç, PAR-2 ˫
'>øɺŲɮ<?!
oe-R‚`kVFe,> GM-CSF -Ȕȓ.NjËʽÍĎį nuclear
factor-kappa B̖NF-κB̗BŸ'ȓ>)ȸ?'>[54]NF-κB .
PAR-2 BŸ'9ǥŲç>)ȸ?'>)<[56]Der f ÚǸ,;
> GM-CSF Ȕȓ,'9üǒ-|LhZ{˫ˋ'>øɺŲ>)
ɮ<?!5!ƒ‡nȏ,> GM-CSF Ȕȓ,.ʽÍĎį nuclear
factor activated T-cell̖NFAT̗-NjËȾʒ;0 Activator protein-1̖AP-1̗
)-ɞû˞ʘ(>)ěā?'=[51]oeR‚`kVFe,'
.NFAT BŸ!WQg„¨˒ɝʻİē'>)9ȸ?'>[2, 3]
47
!$'ɉ‹ɇ(. GM-CSF -Ȕȓ,>WQg„¨˒ɝʻBƬ<,
>)BȮȨ)'PAR-2 BŸ! NFAT -ǥŲç) GM-CSF -Ȕȓ,
%'Ǐʡ>))!
48
2 .5. đ
49
đ 2-1 Der f ;0 E-64 -ǭãę̍>ɚɹţŹ̖48 Ʈ˪Ŧ̗
a) DPBS -6b) Der f 1̖10μg/mL̗c) E-64̖10-5 M̗d) Der f 1̖10μg/mL̗
;0 E-64̖10-5 M̗mˆ. 20μm Bȸ
50
đ 2-2 Der f ;0 E-64 ǭãę̍,>ɚɹȓİƞ̖48 Ʈ˪Ŧ̗
51
đ 2-3 Der f ǭãę̍,> CPEK - GM-CSF Ȕȓ
̜č-ȈɆ!Ķ̐-ŕĖ +S.D.Bȸ̘̠ P < 0.01̖Tukey-Kramer’s
HSD Test̗
52
!
đ 2-4 GM-CSF Ȕȓ,>WYcF‡t†cDˆ\ˮĸÞ-ƃÙå
DŽ
̜č-ȈɆ!Ķ̐-ŕĖ + S.D.Bȸ̘̠ P < 0.01̖Der f -6Bǭã
!ɬBĿǿ)! Dunnet’s Test̗
53
ɉ ̜ɇ
WYcF‡t†cDˆ\BŸ!FiR‚`kVFe,>
GM-CSF-Ȕȓ)NFAT-˫
54
3 .1. Śʫ
R‚`kVFe.ʓȪ-95̕¤ŒBí8>Ǒžɚɹ(=ǒ+VFeLF
‡BȔȓ>),;$'Ȫʀ-ǻȡ:Åȝóů,˫'>[4, 29]R‚
`kVFe,'Ȕȓ?>VFeLF‡-’(9IL-1IL-8CCL17;
0TSLP.D…„OˆŲȪʀȟŴ-ȠŹ,'˞ʘ+VFeLF‡(>
LPS ( ǥ Ų ç ? ! o e - R ‚ ` k V F e . NF-κB ; 0 extracellular
signal-regulated kinase̖ERK̗BŸ'IL-1β;0IL-8BȔȓ>)ȸ
?![70]Fi-ʓȪR‚`kVFeɚɹNJ(>CPEK,'.TNF-α-Ú
Ǹ,;$'p38ƒ‡˜ç?CCL17 mRNA-ʽÍʧŁ?>[111]LƒP
…F‡5BŸ!R‚`kVFe<-TSLP-Ȕȓ,.PAR-2)-Ǧ,İē
>NF-κB-ǥŲç˫'>[13]-;,R‚`kVFe,'
.Ħ˙<-ǒ+ÚǸ,ů'?
?Ȝ+>WQg„¨˒ɝʻǥŲç
?VFeLF‡Ȕȓ?>
Ûɇ,'FiR‚`kVFe<-ȔȓƬ<)+$!GM-CSF9D…
„Oˆ-ÅȝȠŹ,˫>Ôį(>oe-ƒ‡nȏ,'.PMABŸ
!GM-CSF-Ȕȓ,NFAT-ǥŲç˫'>)ȸ?'>[51]˄
ŖoeR‚`kVFe,'9NFATİē[2, 3]WP†KNWSgˆ
\-2-Ȕȓ,˫ˋ'ǥŲç>)Ƭ<,?'>[34]R‚`
kVFe,>GM-CSF-Ȕȓ)NFAT-˫ˋŲ.Ǐʡ?'+
(Ƽɇ(.WYcF‡t†cDˆ\(ǥŲç?!Fi-R‚`kV
Fe- GM-CSFȔȓ,'NFAT˫'>BǏʡ!Ûɇ5(
HDMBȕ'!ĩǾƆÓȄ-ÆƂď˻)+$!)<Ƽȡ(.HDM
üǒ,WYcF‡t†cDˆ\ǥŲBƶ>nnF‡Bȕ!nnF‡.y
GY-Th2ɚɹ;0ǔȇɚɹ-ÔçBµˍ>3[118]PAR-2 BŸ'o
55
e-R‚`kVFeBȯƐǥŲç>)ȸ?'>[100](Ƽɇ
(.nnF‡BŸ!FiR‚`kVFeɚɹNJ̖CPEK̗<-GM-CSFȔȓBELISA,;$'ȷʦŦ„WrH‚ˆ\Da[FBȕ'NFAT-NjË
Ⱦʒ,%'ʣ³!<,WP†Yxƒ‡̖Ciclosporine, CsA̗Bȕ!
ˮĸĶ̐,;$'CPEK-GM-CSFȔȓ,>NFAT-˫-ȿŜ,%'Ǐ
ʡ!
56
3 .2. ƾƢ)ƥǢ
3.2.1. ƾƢ
F i ʓ Ȫ R ‚ ` k V F e - ɚ ɹ NJ ( > CPEK ) - ę ̍ Ǫ CnT-09 . CellnTEC advanced cell system<ʷÆ!5!ǽʑǮęĕOpti-MEM I
̖Invitrogen̗;0ɠ£ę̍Ʈ-ɚɹÝ˺,eƒtW‡ÑȐÞTrypLE express
̖Invitrogen̗B°ȕ!DPBSWYcF‡t†cDˆ\(>nnF‡;
0WYcF‡t†cDˆ\ˮĸÞ(> Trans-Epoxysuccinyl- L-Leucylamido̖4-Guanidino̗Butane̖E-64̗.Sigma-Aldrich<ÆƂ!
3.2.2. ɚɹę̍
̞č<10čɠ£!CPEKB10 mL-CnT-09,Ǩˏ375% CO2ȑĠ(75 cm2r‚YT̖NUNC̗Bȕ'ę̍!80%T‡r„I‡e,˒
!Ǜ˴(ę̍ǪB˰ñ10 mL-DPBS(ɚɹBǤǧ!TrypLE Express
B3 mLã!Ŧ5-10Ô˪ 37,˾ɨɚɹBÝ˺!CnT-09B7mLã
ɕ80̕-ɚɹÝ˺Ǩˏ!)Bȷʦ!ŦɚɹǨˏǪB15 mL
centrifuge tube̖NUNC̗,čôķǰ( 180 G5Ô˪˔Ŭ!˔ŬŦ
ŒǮB˰ñɚɹĝBCnT-09(ÌŽǹ!ÌŽǹ!ɚɹǨˏǪ(2×104 /mL)
B1 mL%24-well plates̖NUNC̗,Ôǣ80̕T‡r„I‡e,˒>5
( 37 5%CO2-ǀ¥(ę̍!-Ŧę̍ǪBOpti-MEM I,ĥƳ
ǽʑǮ(24Ʈ˪ę̍!ŦnnF‡-ǭãBʒ$!DPBS Bȕ'nnF
‡-ő˝ǵǪB̖10-8 M ̤ 10-3 M̗B¯ʕ10 μL BúGH„,ǭã24
Ʈ˪ę̍!t†cDˆ\ˮĸĶ̐(.WYcF‡t†cDˆ\ˮĸÞ(
>E-64Bȕ!DPBS(ǵʞ!E-64ǵʞǪBnnF‡Ǫ,ã'37(30
Ô˪ÑȐ!-ŦnnF‡;0 E-64 ǵʞǪ̖10-8 M ̤ 10-3 M̗5!
57
.DPBSBúGH„-ę̍Ǫˠ-1%̖10 μL̗,+>;,ã37 5%CO2 
(24Ʈ˪ę̍!
3 .2.3. ɚɹȓİȍ
nnF‡;0E-64-ǭãɚɹ-ţŹ;0ȓİȍ,>Ť́Bʣ³>
!8,?
?Bǭã'24Ʈ˪ę̍!ŦɚɹţŹB»Ɇ̇Ūˤ(ʜļ
>))9,Trypan blue 0.4%̖Sigma-Aldrich̗Bȕ'ɚɹȓİȍBǏ
ʡ!
3.2.4. ę̍ŒǮ,>GM-CSFǺŜ
nnF‡5!.E-64)nnF‡-İē(24Ʈ˪ę̍Ŧę̍ŒǮBčô
!"#,-80(Ïɞ·İ!ę̍ŒǮ’-GM-CSFǺŜ. DuoSet canine
GM-CSF ELISA kit̖R&D System̗Bȕ'DZĵ!ǓǴƲɦ.nnF‡
;0E-64Bþ7ǽʑǮę̍Ǫ,;$'ő˝!ƒT‡pg‡eFi-ÿÄŜB
ȕ'¯ž!DZĵĿʮ)+>V‡t„,.nnF‡-^‡nPÔʞǥŲ,
;>ƌƋƅ­-ÔʞBˮĸ>ȮȨ(E-64̖10-3 M̗Bȕˠǜ̚̕)+>;
,ã!
3.2.5. NFAT…xˆ^ˆ˗¨į„WrH‚ˆ\Da[F
ǽʑǮę̍Ǫ(24Ʈ˪ę̍!CPEKBȕ'„WrH‚ˆ\…xˆ^ˆ
D a [ F B Ķ Ʀ ! 50 μL - Opti-MEM I B ȕ ' pGL4-luc2PNFAT-RE-Hygro̖Promega, Madison, AL̗;0pGL4-luc2P- Hygro̖Promega̗
.0.25 μg/well)+>;,pGL4-hRluc-TK̖Promega̗.0.025 μg/well)+
>;,?
?ő˝!?
?-vP^ˆBOpti-MEM 50 μL(ő˝!
Lipofectamine 2000̖Invitorogen̗B 2μL/well)Ǭûķǰ(20Ô˪F‡N
58
€vˆe!Ŧ?Be‚‡YrHPW‡ȕóůǪ)!24-well plate Ë
-ę̍ǪBÿŠDPBS(Ǥǧ!-#Opti-MEM I 0.5 mL/well,›Ɣ'
e‚‡YrHPW‡ȕóůǪB100μL/wellǭã'vP^ˆBŁÆ!̖37
5% CO 24Ʈ˪ę̗̍nnF‡PMA̖50 μM̗;0T‡e†ˆ„)'ő
2
˝!DMSOBǭã'375% CO -ǀ¥(24Ʈ˪ę̍!úwellB
2
DPBS(Ǥǧ!Ŧ,Dual-Luciferase Reporter Assay System̖Promega̗t†eT„,Ǵ'Passive Lysis Buffer 100 μL/wellBã'ķǰ(15Ô˪
Ɗȭ!čô!ǵǪ.!"#,-80(Ïɞ·İ!„zk|ˆ^ˆȕ`€
ˆs̖Sarstedt̗,ü Assay System-LARⅡB 50 μL%ãčô!ǵ
ǪB 10μLBã!Ŧ„zk|ˆ^ˆ̖BioOrbit̗Bȕ'Firefly luciferase
-ȦÄŢŜBDZĵ!5!Stop & Glo Reagent 50 μL9üǒ,ǭãRenilla
luciferase-ȦÄŢŜBDZĵ!ȰĿȨ„WrH‚ˆ\ǥŲ.úV‡t„-ȦÄ
ŢŜǜ̖Firefly luciferase-ȦÄŢŜ/Renilla luciferase-ȦÄŢŜ̗BmockT‡
e†ˆ„-ȦÄŢŜǜ̖Firefly luciferase-ȦÄŢŜ/Renilla luciferase-ȦÄŢ
Ŝ̗(ʔǙ'ɋÓǽǭãɬ)PMA;0nnF‡ǭãɬ(ǜʾ!
3 .2.6. CsA ,;>R‚`kVFe- GM-CSF ȔȓƃÙ-Ǐʡ
̊̎ę̍Ŧę̍ǪB˰ñCsA̖10-6 M5!.10-7 M̗5!. DMSO B
ǭã!ę̍Ǫ,›Ɣ30Ô˪375% CO (ę̍!-Ŧnn
2
F‡5!. PMA B 10 μL(úGH„-ę̍Ǫˠ- 1%) ǭã!24Ʈ˪ę̍Ŧ
ȯ#,ę̍ŒǮBčô-80 (Ïɞ·İ!ę̍ŒǮ’- GM-CSF ǺŜ
.Œ˅!ƥǢ(DZĵ!
3.2.7. ɟʠ
59
ȈɆ!Ķ̐B 3 čĶƦę̍ŒǮ,>GM-CSFǺŜ-ŕĖ¼BɋÓ
!ɟʠʞǃ)'nnF‡ǭãĶ̐,'.nnF‡ǽǭãɬBt†c
Dˆ\ˮĸĶ̐(.E-64Bǭã'+nnF‡ǭãɬB„WrH‚ˆ\
Da[F,%'.DMSO-6Bǭã!ɬB?
?Ŀǿ)! DunnetǏĵ
Bȕ!CsAǭã,;>GM-CSFȔȓƃÙĶ̐,'.nnF‡5!.
PMA,ʧŁ?>GM-CSFȔȓ,Ŀ>NFATNjËȾʒˮĸ,;>ƃÙåDŽB
Ǐʡ>!8Tukey-Kramer-HSDǏĵBȕ'Ç'-ɬ˪Bǜʾ!ɟ
ʠIJȨƶŷŎ. P < 0.05 )ĵɭ!ʞǃ,.ɟʠ]re JMP version 5.1.2
(SAS institute) B°ȕ!
60
3 .3. ɞDŽ
3.3.1. ɚɹ-ţŹ)ȓİȍ
nnF‡̖0.1
20 nM̗;0E-64-İē,'9ɚɹ-ţŹ.ǽǭã
ɬ)Ȱ˓+ɗˢţBȸɚɹ˪;0W~ˆ…ś1-ƐȳŲ9ɡƈ?'
!+<nnF‡ǺŜ 50 nM ¤Œ-Ĝû,'.ɚɹ.ȏţ
,ĥţħƞ-Ǩˏɚɹ9ʦ8<?!̖đ3-1̗nnF‡̖0.1
20 nM̗
;0E-64-?Bǭã!Ĝû,'9ɚɹȓİȍ. 78.7̤83.6% (
$!̖đ3-2̗
3.3.2. nnF‡BŸ!R‚`kVFe,>GM−CSFȔȓ
ę̍24Ʈ˪Ŧ,>ę̍ŒǮ’-ŕĖGM-CSFǺŜ.ǽǭãɬ 12.096
±13.221 pg/mL(nnF‡ 1 nMɬ 55.101 ±12.918 pg/mL5 nMɬ
125.574 ±14.847 pg/mL10 nMɬ 123.603 ±21.033 pg/mL;020 nMɬ
96.312 ±32.009 pg/mL($!nnF‡5nM5(.ȕˠ²İŲ,GM-CSFȔȓ
ˠƶŷ,ġã!̖đ3-3̗
3.3.3. WYcF‡t†cDˆ\ˮĸ,;> GM-CSF Ȕȓ-ƃÙ
nnF‡BŸ!R‚`kVFe-GM-CSFȔȓ.10-10 nM¤Œ-E-64İē
,'ȕˠ,²İ'ƶŷ,ƃÙ?>)Ƭ<)+$!̖đ3-4̗
3.3.4. nnF‡,;> NFAT -NjËȾʒ
nnF‡̖5 nM̗;0PMAǭãɬ,>ȰĿȨ„WrH‚ˆ\ǥŲ.T
‡e†ˆ„ɬ,Ŀ'ƶŷ,̒¼Bȸ!nnF‡ǭãɬ,'.˲Ų
Ŀǿ,Ŀ'ɕ 3.4º-NFAT-NjËȾʒʦ8<?!̖đ3-5̗
61
3 .3.5. CsA,;> GM-CSF Ȕȓ-ƃÙ
CsA-ǭã,;$'PMABŸ!GM-CSF-Ȕȓ.ĴÇ,ƃÙ?!nn
F‡BŸ!GM-CSFȔȓ-ƶŷ+ƃÙ(=+<˙ÔȨ+9-($!̖đ
3-6̗
62
3 .4. ɮļ
nnF‡.WYcF‡t†cDˆ\ǥŲBƶĭ˜ȏ-ɼ̆ɐ[78]ƒ‡
nȏ,>VFeLF‡Ȕȓ[102];0oeǞˑŒȪɚɹ,> TSLP
-ȔȓBµˍ>)ȸ?'>[67]ƼȵɄ(.ÆƂŽøɺ)+$!
ĩǾƆÓė Der f 1 ,£'nnF‡Bȕ!Der f 1 )üǒ, CPEK ,
> GM-CSF -ȔȓBʧŁ!
ʽÍĎį(> NFAT .IL-2 ;0 GM-CSF -ʽÍÙŨ,˞ʘ+ťàB
DŽ!)oe;0yGY-ƒ‡nȏ,'ȸ?'>[119]ʽÍǥŲç-!8,.NFAT ɚɹʶ<NjË1Ⱦʒ>ŭʘ>PMA NFAT -NjËȾʒBµˍ[79]GM-CSF ȔȓBʧŁ>)oe-ƒ‡n
ȏ,'ěā?'>[119]NFAT -NjËȾʒ.L„WG{̙L„}d€
ƒ‡²İŲƒ‡˜çL„Wh€ˆƒ‡,;$'ÙŨ?'>-(NFAT B
Ÿ!ʽÍ.CsA +*,;$'ˮĸ?>[33]ƼȵɄ-ɞDŽ;=nnF
‡ CPEK BǥŲçNFAT -NjËȾʒBʧŁ>)Ƭ<)+$!
+<nnF‡,;$'ʧŁ?> GM-CSF -Ȕȓ.CsA (.ĴÇ
,ƃÙ?˙ÔȨ+ƃÙ($!!$'GM-CSF mRNA -ʽÍ.
NFAT -6+<NF-κB -;+3-WQg„ɝʻ˫'>øɺŲ
ɮ<?>˄ŖCsA Toll-like receptor̖TLR̗2 BŸ' TNF-α -ʽ
ÍBʧŁ!) CPEK ,'ȸ?[43]?. NF-κB -ǥŲç,;>
9-)ɮ<?![55]TLR BŸ! NF-κB -ǥŲç.L„Wh€ˆƒ‡,
;$'ƃÙ?'>)ɮ<?'>[55]<,L„Wh€ˆƒ‡ˮĸ,
;> NF-κB -ǥŲç.myeloid differentiation factor 88 ;0 Toll/IL-1
receptor/resistance domain-containing adaptor-inducing IFN-β̖TRIF̗,²
İ>)yGY-yP†rCˆX,'ȸ?'>[55]oY^z‡
63
BŸ!oe-R‚`kVFe,> GM-CSF -Ȕȓ, NF-κB ˫'
>)ȸ?'>[54]oe-ɸɾɚɹ-ɚɹNJ(> HEK293T ɚɹ
,'.NF-κB . PAR-2 -Ǧ,İē> TRIF BŸ'ǥŲç?>
)ȸ?!PAR-2 .WYcF‡t†cDˆ\,;$'ǥŲç?>)
oe-R‚`kVFe,'ȸ?'>-([88]ƼȵɄ,'ʦ8<?
! CsA ,;> CPEK - GM-CSF Ȕȓ-˙ÔƃÙ.CsA üƮ, TRIF BŸ
' NF-κB BǥŲç!),ʹĎ'>øɺŲ>)ɮ<?!øɺŲBʢƬ>!8,.WYcF‡t†cDˆ\BŸ! NF-κB -ǥŲç
BʢƬ>ŭʘ>
CsA .×£ę̍,>R‚`kVFe-ġǚBƃÙ>)+*ěā
?'>[7]ƼȵɄ-ɞDŽ<NFAT ,ȯƐȨ,¯ȕVFeLF‡
-Ȕȓ9ƃÙ>)ȸ?!
64
3 .5. đ
65
đ 3-1 nnF‡ǭãƮ,>ɚɹţŹ-ĥç̖24 Ʈ˪Ŧ̗
a̗DPBS -6 b̗papain̖20 nM̗ c̗papain̖50 nM̗;0 d̗
papain̖100 nM̗ mˆ. 20 μm Bȸ
66
đ 3-2 nnF‡ǭãę̍,>ɚɹȓİƞ̖24 Ʈ˪Ŧ̗
67
đ 3-3 nnF‡ǭãę̍,> CPEK - GM-CSF Ȕȓ
̜č-ȈɆ!Ķ̐-ŕĖ + S.D. Bȸ̘̠ P < 0.05 ̘̘̠ P < 0.01
̖nnF‡ǽǭãɬBĿǿ)! Dunnet’s Test̗
68
đ 3-4 GM-CSF Ȕȓ,Ŀ>WYcF‡t†cDˆ\ˮĸÞ-ƃÙå
DŽ
̜č-ȈɆ!Ķ̐-ŕĖ + S.D.Bȸ̘̠ P < 0.05 ̘̘̠ P < 0.01
̖nnF‡-6Bǭã!ɬBĿǿ)> Dunnet’s Test̗
69
đ 3-5 nnF‡ǭãŦ- CPEK ,> NFAT NjËȾʒ
̜č-ȈɆ!Ķ̐-ŕĖ+S.D.Bȸ ̘: P < 0.05̘̘̠P < 0.01
(Tukey-Kramer’s HSD Test)
70
đ 3-6 GM-CSF Ȕȓ,> NFAT NjËȾʒˮĸÞ(CsA)-åDŽ
̜č-ȈɆ!Ķ̐-ŕĖ + S.D.Bȸ
̘̠P <0.05̘̘̠ P < 0.01 ̖Tukey-Kramer’s HSD Test̗
71
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72
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78
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