JP2017088559A - フィラグリン産生促進剤 - Google Patents
フィラグリン産生促進剤 Download PDFInfo
- Publication number
- JP2017088559A JP2017088559A JP2015222295A JP2015222295A JP2017088559A JP 2017088559 A JP2017088559 A JP 2017088559A JP 2015222295 A JP2015222295 A JP 2015222295A JP 2015222295 A JP2015222295 A JP 2015222295A JP 2017088559 A JP2017088559 A JP 2017088559A
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- JP
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- Prior art keywords
- filaggrin
- filaggrin production
- acid
- cells
- production promoter
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
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Abstract
Description
項1. 硫酸化グルコサミノグリカンを含有することを特徴とする、フィラグリン産生促進剤。
項2. 前記硫酸化グルコサミノグリカンがヘパリン類似物質である、項1に記載のフィラグリン産生促進剤。
項3. 皮膚外用剤である、項1又は2に記載のフィラグリン産生促進剤。
項4. アトピー性皮膚炎、花粉症、又は尋常性魚鱗癬の予防又は治療に使用される、項1〜3のいずれかに記載のフィラグリン産生促進剤。
硫酸化グルコサミノグリカンとは、アミノ糖を構成単位の1つとして含み、硫酸基を有しているムコ多糖である。
その他の成分
本発明のフィラグリン産生促進剤は、前記硫酸化グルコサミノグリカンの他に、必要に応じて、他の薬理成分を含有していてもよい。このような薬理成分としては、例えば、抗ヒスタミン剤(グリチルリチン酸二カリウム、グリチルリチン酸、ジフェンヒドラミン、塩酸ジフェンヒドラミン等)、局所麻酔剤(プロカイン、テトラカイン、ブピパカイン、メピパカイン、クロロプロカイン、プロパラカイン、メプリルカイン又はこれらの塩、オルソカイン、オキセサゼイン、オキシポリエントキシデカン、ロートエキス、ペルカミンパーゼ、テシットデシチン等)、抗炎症剤(アラントイン、インドメタシン、フェルビナク、ジクロフェナクナトリウム、ロキソプロフェンナトリウム等)、皮膚保護剤(コロジオン、ヒマシ油等)、血行促進成分(ノニル酸ワニリルアミド、ニコチン酸ベンジルエステル、カプサイシン、トウガラシエキス等)、清涼化剤(メントール、カンフル等)、ビタミン類(ビタミンA等)、ムコ多糖類(コンドロイチン硫酸ナトリウム、グルコサミン等)等が挙げられる。
本発明のフィラグリン産生促進剤は、皮膚外用剤、内服剤等のいずれの剤型であってもよいが、フィラグリンの産生をより一層効果的に促進させるという観点から、好ましくは皮膚外用剤が挙げられる。
本発明のフィラグリン産生促進剤は、表皮細胞(特に、表皮の上層に存在する細胞)においてフィラグリンの前駆体であるプロフィラグリンmRNAの発現量を増大させることにより、フィラグリンの産生を促進することができる。従って、本発明のフィラグリン産生促進剤は、フィラグリンの機能低下が発症要因の一つになっている疾患や症状の予防又は治療に有効である。フィラグリンの機能低下が発症要因の一つになっている疾患や症状としては、具体的には、アトピー性皮膚炎、喘息、鼻炎、花粉症、金属アレルギー等のアレルギー疾患、尋常性魚鱗癬、ウイルスや細菌による皮膚感染症等が挙げられる。
硫酸化グルコサミノグリカンが表皮細胞のフィラグリン産生に与える影響を評価するために以下の実験を行った。
(細胞)
本試験例において細胞は以下の2種類を使用した。
(A)通常表皮モデル細胞(HaCaT細胞)
通常表皮モデル細胞として、HaCaT細胞(ヒト表皮角化細胞(Deutsches Krebsforschungszentrum,Stiftung desoffentlichen Rechts(DKFZ),Heidelberg,Germany))を使用した。
(B)表皮上層モデル細胞(シンタキシン4高発現HaCaT細胞)
ある程度分化が進んだ表皮上層部の細胞モデルを作製し、試験に使用した。(i)表皮細胞は角化すると基底層から有棘層・顆粒層に移動すること、(ii)上層に移動するに従い、細胞外にシンタキシン4が多く分泌されること、及び(iii)HaCaT細胞においてシンタキシン4を高発現させると角化が進行すること(Kadono N.et al.,Mol Med 2015)が知られている。そこで、表皮上層モデル細胞として、シンタキシン4が高発現しているHaCaT細胞を作製し、本試験例において使用した。作製方法は、以下の通りである。
先ず、細胞外に強制的に提示させるために21アミノ酸残基からなるマウス由来IL−2シグナルペプチドをコードする遺伝子をマウスシンタキシン4遺伝子のN末端に付加し、これをpQCXINレトロウイルスベクターのマルチクローニングサイトに挿入した。次に、これをトランスフェクション試薬Lipofectamine PLUS(サーモフィッシャーサイエンティフィック株式会社製)を用いて、パッケージング細胞PT67に導入した。得られた細胞の培養上清からレトロウイルスを得た。得られたウイルスをHaCaT細胞に感染させることにより、細胞外にシンタキシン4を強制発現したHaCaT細胞(表皮上層モデル細胞)を作製した。
D−MEM/Ham’s F−12(和光純薬工業株式会社)/10%FCS添加
ヘパリン類似物質
(1)通常表皮モデル細胞におけるプロフィラグリンmRNAの発現量
12ウェルプレートの各ウェルにモデル細胞を1×105個/ウェルの濃度でそれぞれ播種し、37℃、5%CO2条件下で培養した。翌日に、各ウェルにヘパリン類似物質を0.5mg/mLとなるように添加し、37℃、5%CO2条件下で培養した。その翌日に、モデル細胞を回収しRNeasy mini kit(株式会社キアゲン製)を用いて全RNAを抽出した。その後、ReverTra Ace(東洋紡株式会社製)を用いて逆転写を行い、cDNAを合成した。次いで、Fast Start Essential DNA Green Master on LightCycler Nano system(ロシュ・ダイアグノスティックス株式会社製)を用いて、qRT−PCRにてプロフィラグリン遺伝子の増幅を行った。β−アクチンを内部標準として使用し、プロフィラグリンmRNAの発現量を標準化した。このmRNAの発現量を、ヘパリン類似物質を添加しなかった以外は前記と同様にして確認したmRNAの発現量(コントロール)で除算した。
通常表皮モデル細胞に代えて表皮上層モデル細胞を使用した以外は、前記(1)と同様に操作し、プロフィラグリンmRNAの発現量を確認し、コントロール(ヘパリン類似物質を添加しなかった場合のmRNA発現量)で除算した。
表1〜2に示す組成のクリーム剤(処方例1〜8)、表3に示すローション剤(処方例9〜14)、表4に示すジェル剤(処方例15〜19)、及び表5〜6に示す乳液剤(処方例20〜29)を調製した。これらの製剤は、いずれも、前記試験例1の場合と同様に、表皮細胞のプロフィラグリンmRNAの発現量を増大させる効果が期待され、フィラグリンの産生促進に有効である。
Claims (4)
- 硫酸化グルコサミノグリカンを含有することを特徴とする、フィラグリン産生促進剤。
- 前記硫酸化グルコサミノグリカンがヘパリン類似物質である、請求項1に記載のフィラグリン産生促進剤。
- 皮膚外用剤である、請求項1又は2に記載のフィラグリン産生促進剤。
- アトピー性皮膚炎、花粉症、又は尋常性魚鱗癬の予防又は治療に使用される、請求項1〜3のいずれかに記載のフィラグリン産生促進剤。
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