新潟大学Niigata University博士（医学）新大院博(医)第1045号doctoral thesi

Nguyen, Hong Ha

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博 士 論 文 の 要 旨 及 び 審 査 結 果 の 要 旨  氏       名   NGUYEN Hong Ha 学       位   博士（医学）                           学 位 記 番 号    新大院博（医）第 1045 号 学位授与の日付   令和 4年3月23日 学位授与の要件   学位規則第 4条第1項該当 博 士 論 文 名   New insight of itch mediators and proinflammatory cytokines in epidermolysis bullosa. (表皮水疱症におけるかゆみメディエーターと炎症性サイトカインの新しい洞察)            論文審査委員    主査 教授 松田 健 副査 教授 平島 正則 副査 講師 濱 菜摘    博士論文の要旨                                Introduction: Epidermolysis bullosa (EB) is a hereditary disorder characterized by mechanical stress-induced blistering. The presence of extracutaneous complications such as cardiomyopathy and renal disease observed in severe EB subtypes and the fact that pruritus is a common symptom across all EB subtypes indicate that EB is not only a skin fragility disease but also a systemic inflammatory disorder. This study aims to elucidate the basis of the systemic inflammation seen in EB patients. Methods: The serum samples of 20 EB patients were analyzed by Luminex cytokine assays and enzyme-linked immunosorbent assays. Results: The serum levels of sIL-2R, IL-6, HGF, M-CSF, SCGF-β, IL-8, IL-16, IFN-γ, MIF, MIP-1α, and TSLP (p < 0.01, p < 0.01, p < 0.01, p < 0.01, p < 0.01, p < 0.05, p < 0.05, p < 0.05, p < 0.05, p < 0.05, and p = 0.01, respectively) were found to be significantly elevated in EB patients, whereas TNF-β (p < 0.01) was decreased. Th2 cytokines including IL-4, IL-5, and IL-13 were not elevated in EB patients. In contrast, TSLP was significantly increased, and IL-31 and oncostatin M were not statistically significant but tended to be higher in the patients than in healthy controls. Among proinflammatory cytokines such as IL-1β, IL-6, and TNF-α, IL-6 was elevated in EB patients. Discussion: In this study, Th2 cytokines such as IL-4, IL-5, and IL-13 were investigated and, in agreement with previous reports, Th2 cytokines in the sera of EB patients were not significantly different from those of healthy individuals. On the other hand, TSLP in EB was significantly higher than in healthy controls. Although the results of this study did not show a trend towards higher TSLP levels in patients with higher affected body surface areas, it may be that TSLP is more likely to be produced by epidermal keratinocytes in patients with EB who often have chronic wounds prone to infection and cutaneous dysbiosis. In addition, there was a tendency for IL-31, known as a pruritogen, to be elevated in DEB patients compared to healthy controls. The expression of the IL-31 gene is upregulated in itchy human skin diseases such as atopic dermatitis (AD), allergic contact dermatitis, psoriasis, and prurigo nodularis. Furthermore, serum levels of IL-31 are elevated in AD, and IL-31 levels correlate with disease severity, suggesting that IL-31 plays a pivotal role in the induction and maintenance of pruritus and chronic skin inflammation in AD. Recently, nemolizumab, the humanized anti-human IL-31 receptor A (IL-13RA) monoclonal antibody, has been reported to improve pruritus in atopic dermatitis and prurigo nodularis, and IL-31 may also provide a new target for anti-pruritic and anti-inflammatory activity in the treatment for EB. The proinflammatory cytokines, including IL-1β, IL-6, and TNF-α, from the serum of EB, patients were analyzed. In the present study, serum IL-6 levels were significantly elevated in patients with DEB. Consistent with these reports, in two previous studies, serum IL-6 levels were significantly elevated in patients with EB and correlated with the severity of the disease. These facts suggest that IL-6 can play a pathophysiological role in triggering serological inflammation in patients with EB. Conclusions: The imbalance of several itch mediators and proinflammatory cytokines was identified. In the future, biologics that target the cytokines found to be elevated in the sera of patients may be a beneficial treatment option for EB.  審査結果の要旨 20 名の表皮水疱症患者血清中の炎症性サイトカイン、痒み関連のサイトカイン（IL-2R, IL-6, HGF, M-CSF, SCGF-β, IL-8, IL-16, IFN-γ, MIF, MIP-1α, TSLPなど）をLuminex bead-based cytokinesアッセイおよびELISAキットを用いてスクリーニング、測定した結果、炎症性サイトカイン IL-6においては健常コントロールより高値を認めた。痒み関連のサイトカイン IL-4, IL-5, IL-13に関しては健常コントロールと有意差はなく、これは従来報告されているとおりであったが、IL-31とTSLPに関しては高値であった。 この結果に基づき、TSLP, IL-31, IL-6に関しては表皮水疱症の治療の新たなターゲットとなる可能性を有していると考えられる。 また、複合ヘテロ接合型変異を有した表皮水疱症モデルマウスに対し、遺伝子編集技術によって遺伝子変換を誘導した。具体的には AAVを用いて原因遺伝子に特異的な CRISPR/Cas9をモデルマウス由来の表皮角化細胞およびin vivoに直接遺伝子導入し、タンパク質の発現が認められるようになった。本研究では復帰変異モザイクの発症機序の解明のみならず、新たな遺伝子治療法につながる概念実証を行った。 本論文は、表皮水疱症患者の血清中のサイトカイン濃度の詳細な解析により新たな治療ターゲットとなるサイトカインを見出し、また表皮水疱症モデルマウスをはじめて報告し、将来的な表皮水疱症の遺伝子治療実現の可能性を示したことに学位論文としての価値を認める。 

表皮水疱症におけるかゆみメディエーターと炎症性サイトカインの新しい洞察

https://niigata-u.repo.nii.ac.jp/record/2000831/files/r3nmk1045_a.pdf

表皮水疱症におけるかゆみメディエーターと炎症性サイトカインの新しい洞察

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