選擇性誘導XBP1s 藥物--IXA4對腎臟的保護作用

Abstract

急性腎損傷（AKI）的定義為在短時間內腎功能急遽惡化。雖然腎臟有適應性修復的能力，但嚴重的損傷也可能引發異常的修復，造成不可逆轉的損害，最終導致慢性腎臟病（CKD）和末期腎臟病（ESRD）。最近的研究揭示了導致從AKI過渡到CKD的異常修復的各種機制。例如腎小管上皮細胞因為停滯於細胞週期之G2/M期，進而增強纖維化因子的表達。

內質網壓力活化與腎臟疾病進展密切相關。當發生內質網壓力時，未摺疊蛋白反應（Unfolded protein response, UPR）會啟動來維護內質網蛋白質穩態。未摺疊蛋白反應的不同途徑中，肌醇需求酶1α（Inositol-requiring enzyme 1α, IRE1α）的磷酸化會促進X-box結合蛋白（X-box binding protein, XBP1）RNA截切成splicing form XBP1 (XBP1s)，促進適應性未摺疊蛋白反應，進而增強蛋白質折疊能力和內質網蛋白質降解。

先前研究已證實適應性UPR調節因子-XBP1s在急性腎損傷至慢性腎臟病轉變中扮演關鍵角色，當XBP1s表現缺損下，將造成更嚴重的腎損傷後纖維化進展；此外，XBP1基因敲除會導致受損的腎小管上皮細胞不完全修復。基於以上研究基礎，提出XBP1s選擇性活化可能作為腎損傷後治療策略之假說。

為了驗證該假說，本研究將利用選擇性XBP1s激活劑-IXA4以評估腎損傷後之治療效果及其詳細機轉。於細胞實驗中使用人類腎近曲小管上皮細胞（HK-2），透過TGF-β1模擬腎臟纖維化過程之微環境，並同時給予IXA4。結果顯示IXA4具有腎小管上皮細胞的保護效果，透過抑制由TGF-β1所引起的促纖維化、上皮間質轉化、促炎症以及G2/M細胞週期停滯。在動物實驗中，本研究使用單側缺血再灌流損傷和單側尿路阻塞的動物模型進行測試，以腹腔注射的方法施打IXA4。結果顯示，於單側缺血再灌注損傷的動物模型中，IXA4顯著減少了結締組織生長因子（CTGF）蛋白表現；於單側尿路阻塞的情況下，IXA4治療則無明顯改善纖維化之趨勢。

總而言之，IXA4活化XBP1剪接減輕由TGF-β引起的促纖維化過程、EMT和G2/M細胞週期停滯。在體內實驗中，IXA4在減緩腎臟進展方面有潛在作用，然而後續仍需要進一步研究探討IXA4在複雜的體內環境中的有效性。

Acute kidney injury (AKI) manifests as a swift deterioration in renal function within a brief timeframe. Although kidneys can adaptively repair functional losses, severe injuries may trigger maladaptive repair, causing irreversible damage and culminating in chronic kidney disease (CKD) and end-stage renal disease (ESRD). Recent research has uncovered various mechanisms of maladaptive repair that contribute to the transition from AKI to CKD. Mechanisms such as G2/M arrest of tubular epithelial cells, which results in heightened expression of pro-fibrotic factors.

Endoplasmic reticulum (ER) stress has been identified as a mediator of kidney disease progression. Upon ER stress, the unfolded protein responses (UPRs) are activated to maintain ER proteostasis. Among the different pathways of UPRs, inositol-requiring enzyme 1α (IRE1α)-mediated alternative splicing of X-box binding protein (XBP1) promotes adaptive UPRs by enhancing protein folding capacity and ER-associated protein degradation. Our recent work has demonstrated that the down-regulation of XBP1 promotes the transition from AKI to CKD. To translate this concept into clinical application, we applied IXA4, a selective XBP1s activator, in preclinical study.

Human kidney proximal tubular epithelial cells (HK-2) were treated with TGF-β to simulate the microenvironment of fibrotic kidneys, and IXA4 was co-administered for 48 hours. In animal models of unilateral ischemia-reperfusion injury (uIRI) and unilateral ureteral obstruction (UUO), mice received IXA4 treatment through intraperitoneal (ip) injections once daily during the specified time points.

In vitro experiments using HK-2 cells demonstrated that IXA4 treatment had positive effects. It reduced pro-fibrotic processes, epithelial-mesenchymal transition (EMT), pro-inflammation, and G2/M cell cycle arrest induced by TGF-β, a key factor in kidney disease progression. IXA4 was then tested in an AKI to CKD animal model-uIRI. IXA4 significantly decreased connective tissue growth factor (CTGF) protein expression, suggesting a potential reduction in fibrosis. In the case of UUO, IXA4 treatment had no effect in ameliorating renal fibrosis and EMT process.

XBP1 splicing promoted by IXA4 alleviated pro-fibrotic processes, EMT, and G2/M cell cycle arrest induced by TGF-β in vitro. Furthermore, when tested in vivo, IXA4 appeared to play a potential role in retarding uIRI caused kidney disease progression. These findings suggest that further research is required to fully understand the effectiveness of IXA4 in the complex in vivo environment.