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重症喘息における好酸球性炎症制御に関する研究

板倉 康司 東北大学

2020.03.25

概要

背景
気管⽀喘息 (以下、喘息) は、成⼈の約10%が罹患する国⺠病の⼀つである。喘息のうち 5-15%は既存の治療薬を使⽤しても症状のコントロールが困難な重症喘息であり、重症喘 息の⽣活の質や⽣命予後を改善させるために新規治療法の開発が急務である。気道上⽪細胞は、pathogen-associated molecular patterns (PAMPs) やdamage-associated molecular patterns (DAMPs) を認識することでサイトカイン、ケモカインを産⽣し、2型炎症を惹起する起点となる。しかし、気道上⽪細胞によって誘導される2型炎症がどのように抑制されているかについては不明な点が多い。TAM (Tyro3、Axl、Mertk) 受容体ファミリーのうちの1つであるAxl 受容体チロシンキナーゼは、肺胞マクロファージや樹状細胞に発現し、アポトーシス細胞の貪⾷による efferocytosis の促進や Toll-like receptor (TLR) シグナルの抑制を介して炎症の収束に寄与する。私は、Axl が気道上⽪基底細胞に発現し、インフルエンザウイルス感染による傷害後の気道上⽪の細胞増殖と組織修復を促進させることを報告した。しかし、Axl が気道上⽪細胞を介した2型炎症にどのように関与しているかは不明である。そこで、本研究では2型気道炎症における Axl の役割を明らかにすることを⽬的とした。

⽅法
喘息の気道上⽪における Axl 発現の変化を検討するため、⼆つの独⽴したコホートを⽤いて、健常者、軽症・中等症喘息、重症喘息の間でヒト気道上⽪細胞の遺伝⼦発現を解析した。喘息患者の気管⽀⽣検組織の免疫染⾊により、⾮喘息、軽症・中等症喘息、重症喘息の間で気道上⽪細胞におけるAxl 蛋⽩の発現を⽐較した。喘息患者の気道上⽪細胞におけるAxl 蛋⽩の発現と、気道組織に浸潤する好酸球数とマスト細胞数の関連について病理組織学的に検討した。2型気道炎症におけるAxl の機能を解析するため、house dust mite 抽出物 (HDM) をマウスへ経⿐投与し2型気道炎症モデルを作成し、HDM 投与後のAxl−/−マウス気管、肺組織に浸潤する炎症細胞数をフローサイトメトリーにより定量した。ヒト気道上⽪細胞株 (BEAS-2B 細胞) を⽤いて、small interfering RNA (siRNA) によりAxl をノックダウンし、Axl が制御する気道上⽪細胞由来のサイトカイン、ケモカインを検討した。

結果
ヒト気道上⽪細胞の遺伝⼦発現解析では、⼆つのコホートの両⽅において、重症喘息の気道上⽪細胞におけるAXL mRNA の発現が有意に低下していた。重症喘息の気管⽀⽣検組織では、気道上⽪細胞におけるAxl 蛋⽩の発現が有意に低下していた。全⾝性副腎⽪質ステロイド投与後の喘息患者の気管⽀組織に浸潤する好酸球数とマスト細胞数は、いずれも気道上⽪細胞の Axl 蛋⽩の発現と強い負の相関を⽰した。これらの結果から、気道上⽪細胞における Axl の発現低下は、重症喘息における2型気道炎症およびステロイド抵抗性の病態と関連している可能性が⽰唆された。In vivo における検証では、HDM 投与群 Axl−/−マウス肺組織に浸潤する好酸球数は HDM 投与群野⽣型マウスと⽐較し有意な差を認めなかった。⼀⽅、HDM 投与群Axl−/−マウス気管組織に浸潤する好酸球数は、HDM ⾮投与群Axl−/−マウスおよびHDM 投与群野⽣型マウスと⽐較して有意に増加した。これらの結果により、Axl は2型気道炎症において好酸球浸潤を抑制していることが明らかとなった。BEAS-2B 細胞を⽤いた in vitro の検討では、Axl のノックダウンにより、HDM 刺激後における granulocyte macrophage colony-stimulating factor (GM-CSF)、regulated on activation, normal T cell expressed and secreted (RANTES) の遺伝⼦および蛋⽩の発現が有意に上昇した。更に、GM-CSF とRANTESの発現を促進させる転写因⼦ nuclear factor 𝜅 B (NF𝜅B) の活性化阻害因⼦である phosphorylated A20の発現を検討したところ、Axl のノックダウンによって HDM 刺激後における phosphorylated A20の発現は低下した。これらの結果により、気道上⽪細胞に発現する Axl は HDM 刺激時において、A20のリン酸化を促進することで NF𝜅B の活性化を阻害し、 GM-CSF とRANTES の遺伝⼦発現を抑制する可能性が⽰唆された。

結論
本研究は、Axl が好酸球浸潤を抑制することで2型気道炎症を負に制御することを明らかにした。また、Axl は気道上⽪細胞において HDM 刺激時にA20のリン酸化を誘導し、GM-CSFと RANTES の遺伝⼦発現を抑制することで、好酸球の活性化と組織への浸潤を阻害する可能性が⽰唆された。更に、気道上⽪細胞における Axl の発現低下が、重症喘息におけるステロイド抵抗性の2型気道炎症に関与している可能性があることを⽰した。本研究を発展させることにより、重症喘息の病態解明が進歩し、重症喘息の新規バイオマーカーおよび新規治療法の開発が進展することが期待される。

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