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メタボロミクスを活用した肝細胞における中鎖脂肪酸代謝物解析

伏見, 達也 FUSHIMI, Tatsuya フシミ, タツヤ 九州大学

2021.03.24

概要

炭素鎖が 6-12 の脂肪酸は広義には中鎖脂肪酸 (MCFAs) と呼ばれ,ココナッツ油やパーム核油中に,トリアシルグリセロール (TG) の形で主に中鎖脂肪酸油 (MCTs) として存在している.MCTsは長鎖脂肪酸油 (LCTs) と比較して,物性面に限らず,摂取時の消化・吸収・代謝の面においても大きく異なる性質を持つ.例えば,MCFAs は肝臓にてグルコース代替のエネルギー源となるケトン体を産生すると考えられていることから,様々な臨床研究領域で注目されている.ケトン体産生の場である肝臓を対象とした MCFAs の代謝研究は,主に肝細胞を用いた in vitro で実施されてきた.しかしながら,従来の研究では MCFAs の肝代謝の一部についての報告に留まっており,代謝総体の定量的な機能解析はなされておらず,MCFAs 代謝の全容,代謝運命に関しては十分な解明には至っていない.また,個々の鎖長の異なる MCFA の代謝特性についても全てが明らかにされているわけではない.そこで,本研究では,肝細胞である AML12 細胞を用いて,MCFAs および長鎖脂肪酸(LCFAs) で処理した AML12 細胞のメタボローム解析を行った.種々のメタボローム分析技術を駆使したメタボリックプロファイリングおよび安定同位体標識 MCFAを使用した動的メタボローム解析を行うことで,MCFAs の代謝特性,代謝運命を明らかにすることを目的とした.

第二章では,AML12 細胞における MCFA/LCFA 間,MCFA 種間の代謝評価を実施するために,測定対象となる代謝物の物理的特性に応じて,3 種の親水性代謝物分析法と 2 種の疎水性代謝物分析法を用いたワイドターゲットメタボローム分析を実施した.その結果,各脂肪酸で処理した AML12細胞から 183 種の親水性代謝物および 688 種の疎水性代謝物の計 871 種の代謝物を同定,相対定量することに成功した.得られた代謝プロファイルデータから,ラウリン酸 (FA 12:0) 処理は,オクタン酸 (FA 8:0) およびデカン酸 (FA 10:0) と比較して代謝プロファイルが大きく異なり,TG,コレステロール (Cholsterol),コレステロールエステル (ChE),ホスファチジルエタノールアミン (PE),リゾホスファチジルエタノールアミン (LPE),リゾホスファチジルコリン (LPC),セラミド (Cer),ヘキソシルセラミド (HexCer) の各脂質量を増加させることを明らかにした.一方,FA 8:0 およびFA 10:0 による処理は AML12 細胞のケトン体産生が亢進した.また,FA 8:0 はコントロールと比較して TG 組成を変化させるものの,TG 量並びにその他脂質量においても有意な差は見られなかった.LCFAs に分類されるオレイン酸 (FA 18:1) による処理は TG 合成経路に関与するホスファチジン酸(PA),ジアシルグリセロール (DG),TG 量を増加させ,TG 合成の基質として利用されることが示唆された.

第三章では,メタボローム分析技術と安定同位体標識を組み合わせた代謝ターンオーバー解析を活用し,AML12 細胞を 13C8-FA 8:0 および 13C18-FA 18:1 で処理することで FA の代謝動態を観察した.その結果,FA 8:0 はアセチル-CoA に迅速に変換されケトン体,クエン酸回路中間体,一部の糖原生アミノ酸に資化されることを定量的に示した.一方の FA 18:1 は第二章で示唆された TG 合成の基質として資化されることを TG の 13C 標識化率を指標として初めて示した.

本博士論文では,MCFA/LCFA 間,MCFA 種間の包括的かつ定量的な代謝評価によって,肝細胞における MCFAs の代謝特性,代謝運命を明らかにした.今後,動物モデルや他の細胞種を対象としたメタボローム分析による MCFAs の新たな代謝特性や機能の発見が,栄養状態や疾患に応じた油脂の提供の一助となり,油脂を通じた健康課題の改善に繋がると期待される.

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参考文献

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