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標題: | 內質網蛋白TXNDC5穩定肺纖維母細胞TGFBR1蛋白表現以促進TGFβ訊息傳遞進而引起肺纖維化 Endoplasmic Reticulum Protein TXNDC5 Promotes Pulmonary Fibrosis by Augmenting TGFβ Signaling via Post-translational Stabilization of TGFBR1 in Lung Fibroblasts |
作者: | Tzu-Han Lee 李姿涵 |
指導教授: | 楊鎧鍵(Kai-Chien Yang) |
關鍵字: | 肺纖維化,肺纖維母細胞,肺功能, pulmonary fibrosisoblast,TXNDC5,lung function,TGFBR1,pulmonary fibroblast, |
出版年 : | 2020 |
學位: | 博士 |
摘要: | 肺纖維化可以是原發性 (idiopathic pulmonary fibrosis, IPF)或次發於其他肺部疾病,如急性呼吸窘迫症候群 (acute respiratory distress syndrome, ARDS)及嚴重特殊傳染性肺炎 (COVID-19),或其他系統性疾病,如多發性硬化症 (multiple sclerosis)及類風溼性關節炎 (rheumatoid arthritis)。肺纖維化會破壞肺泡組織結構,於間質產生嚴重結痂 (scar),導致肺泡氣體交換功能受阻及肺活量下降,進而造成血氧降低、呼吸困難及運動耐受力降低的症狀。除了呼吸系統的症狀,肺纖維化也會導致肺動脈高壓、右心衰竭,以及肺栓塞風險的增加。由於肺纖維化引起的健康影響與死亡率十分嚴重,病人被診斷為 IPF後其中位存活時間 (medium survival time)只有二到四年,盛行率也隨人口老化逐 漸攀升,是醫療與公衛上亟待解決的課題。然而,目前針對肺纖維化的治療選擇非常有 限,需要更多研究來找尋引發肺纖維化的新型因子及致病機轉,來發展治療肺纖維化的新型藥物。 我們的研究團隊先前發現一個內質網蛋白 Thioredoxin Domain Containing 5 (TXNDC5)在心臟纖維化的過程扮演重要角色。我們發現 TXNDC5在纖維化的心臟組織中表達量顯著上升,且能透過兩種不同機轉促進心肌纖維化:其一能透過增加纖維母細胞中氧化壓力,激活 JNK而使纖維母細胞活化增生,其二是能透過其protein disulfide isomerase活性來協助 ECM蛋白折疊,增加 ECM分泌的產 量。我們在小鼠進行 Txndc5基因剔除後,發現心臟受 beta-agonist刺激造成的心肌功能受損及心肌纖維化都能獲得改善。透過生物資訊學的分析,我們發現 TXNDC5在 IPF病患的肺組織中高度增加,其表現量亦與其他肺纖維化相關蛋白呈高度正相關。我們推測,TXNDC5在肺纖維化可能也具有重要角色。我們的實驗結果發現在肺纖維化可能也具有重要角色。我們的實驗結果發現IPF病病患肺組織及肺纖維母細胞中的患肺組織及肺纖維母細胞中的TXNDC5基因及蛋白表達量比正常人高。基因及蛋白表達量比正常人高。TXNDC5高度且特定表現在高度且特定表現在bleomycin誘導之小鼠肺纖維化模型中的肺纖維母細胞。誘導之小鼠肺纖維化模型中的肺纖維母細胞。TXNDC5主要透過影響肺纖維母細胞中主要透過影響肺纖維母細胞中TGFBR1的穩定性及表現量來強化的穩定性及表現量來強化TGFβ訊息傳遞路徑,造成肺纖維母細胞的大量活化增生及胞外基質堆積,引起肺纖維化。訊息傳遞路徑,造成肺纖維母細胞的大量活化增生及胞外基質堆積,引起肺纖維化。在小鼠的肺纖維化動物模型中也證實,利用誘導型在小鼠的肺纖維化動物模型中也證實,利用誘導型CRISPR/Cas9基因編輯技術敲基因編輯技術敲除纖維母細胞中除纖維母細胞中Txndc5基因表現,能有效減緩肺纖維化之進程並改善肺功能。綜基因表現,能有效減緩肺纖維化之進程並改善肺功能。綜合以上結果顯示,抑制TXNDC5活性可望能作為肺纖維化病患的新型治療藥物。 RATIONALE: Pulmonary fibrosis (PF) has become a major and growing public health problem. PF can be idiopathic or secondary to various diseases such as acute respiratory distress syndrome and COVID-19. PF can distort pulmonary architecture and cause excessive scarring which leading to impaired lung function and alveolar gas exchange, finally resulting in hypoxemia, dyspnea and exercise intolerance. In spite of the advances in medicine, the incidence and mortality rate of PF remain high and options for PF therapeutics are very limited. There is a clear need to identify novel mediators and pathways of lung fibrosis to develop therapies to improve the outcomes of PF patients. Recently, our group has identified thioredoxin domain containing 5 (TXNDC5), a cardiac fibroblast (CF)-enriched endoplasmic reticulum (ER) protein, as a novel mediator of cardiac fibrosis. Through the bioinformatic approach, we have found that TXNDC5 was highly up-regulated and positively correlate with extracellular matrix (ECM) proteins in lung tissues from patients with idiopathic pulmonary fibrosis (IPF). Based on these findings, we hypothesize that TXNDC5 could also contribute to the development of lung fibrosis. METHODS: To investigate the role of TXNDC5 in vivo, WT, Txndc5-/- and fibroblasts-specific Txndc5 deletion (Col1a2-Cre/ERT2*Txndc5fl/fl) mice were subjected to intratracheal bleomycin (BLM, 3mg/kg) treatment. The extent and functional consequences of pulmonary fibrosis were evaluated using histological study, immunofluorescence, pulmonary function test, and transcript analysis. To determine the molecular mechanisms by which TXNDC5 regulates lung fibroblasts in vitro, lentiviral vectors was used to knockdown or overexpress TXNDC5 in primary human pulmonary fibroblasts (HPF-a). Primary mouse lung fibroblasts isolated from WT and Txndc5-/- mice were used for mechanistic investigations. RESULTS: Here we show that TXNDC5 is highly upregulated in the lung fibroblasts from both patients with idiopathic pulmonary fibrosis and a mouse model of bleomycin (BLM)-induced PF. Global deletion of Txndc5 significantly reduces the extent of PF and preserves lung function in mice treated with BLM. Mechanistic investigations demonstrate that TXNDC5 promotes fibrogenesis by enhancing TGF 1 canonical and non-canonical signaling through direct binding with and stabilization of TGFBR1 in lung fibroblasts. Moreover, TGF 1 stimulation is shown to upregulate TXNDC5 via ER stress/ATF6-dependent transcriptional control in lung fibroblasts. Furthermore, inducing fibroblast-specific deletion of Txndc5 lessons the progression of BLM-induced PF and lung function deterioration. CONCLUSIONS: The present study revealed a novel role for TXNDC5 in the development of PF. TXNDC5 increases in fibrotic lungs and promotes pulmonary fibroblast activation, proliferation and ECM production by enhancing TGFBR1 expression and stabilization. Targeted deletion of Txndc5 protects against bleomycin-induced PF and lung dysfunction. Therefore, targeting TXNDC5 could be a novel therapeutic approach against PF. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/72971 |
DOI: | 10.6342/NTU202004476 |
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顯示於系所單位: | 藥理學科所 |
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