Liver is a unique organ in displaying a reparative and regenerative response after acute/chronic damage or partial hepatectomy, when all the cell types must proliferate to re-establish the liver mass. The NADPH oxidase NOX4 mediates Transforming Growth Factor-beta (TGF-β) actions, including apoptosis in hepatocytes and activation of stellate cells to myofibroblasts. Aim of this work was to analyze the impact of NOX4 in liver regeneration by using two mouse models where Nox4 was deleted: 1) general deletion of Nox4 (NOX4−/−) and 2) hepatocyte-specific deletion of Nox4 (NOX4hepKO). Liver regeneration was analyzed after 2/3 partial hepatectomy (PH). Results indicated an earlier recovery of the liver-to-body weight ratio in both NOX4−/− and NOX4hepKO mice and an increased survival, when compared to corresponding WT mice. The regenerative hepatocellular fat accumulation and the parenchyma organization recovered faster in NOX4 deleted livers. Hepatocyte proliferation, analyzed by Ki67 and phospho-Histone3 immunohistochemistry, was accelerated and increased in NOX4 deleted mice, coincident with an earlier and increased Myc expression. Primary hepatocytes isolated from NOX4 deleted mice showed higher proliferative capacity and increased expression of Myc and different cyclins in response to serum. Transcriptomic analysis through RNA-seq revealed significant changes after PH in NOX4−/− mice and support a relevant role for Myc in a node of regulation of proliferation-related genes. Interestingly, RNA-seq also revealed changes in the expression of genes related to activation of the TGF-β pathway. In fact, levels of active TGF-β1, phosphorylation of Smads and levels of its target p21 were lower at 24 h in NOX4 deleted mice. Nox4 did not appear to be essential for the termination of liver regeneration in vivo, neither for the in vitro hepatocyte response to TGF-β1 in terms of growth inhibition, which suggest its potential as therapeutic target to improve liver regeneration, without adverse effects.

肝脏是一种独特的器官，在急性/慢性损伤或部分肝切除术后表现出修复和再生反应，此时所有细胞类型都必须增殖以重建肝脏肿块。NADPH 氧化酶 NOX4 介导转化生长因子-β (TGF-β) 作用，包括肝细胞凋亡和星状细胞活化为肌成纤维细胞。这项工作的目的是通过使用两种缺失Nox4的小鼠模型分析 NOX4 在肝再生中的影响：1） Nox4的一般缺失（NOX4-/-）和 2） Nox4的肝细胞特异性缺失(NOX4hepKO)。在 2/3 部分肝切除 (PH) 后分析肝再生。结果表明，与相应的 WT 小鼠相比，NOX4-/- 和 NOX4hepKO 小鼠的肝脏与体重比较早恢复，存活率增加。在 NOX4 缺失的肝脏中，再生性肝细胞脂肪积累和实质组织恢复得更快。通过 Ki67 和磷酸化组蛋白 3 免疫组织化学分析，NOX4 缺失小鼠的肝细胞增殖加速和增加，与早期和增加的Myc表达一致。从 NOX4 缺失小鼠中分离的原代肝细胞显示出更高的增殖能力和Myc的表达增加以及响应血清的不同细胞周期蛋白。通过 RNA-seq 进行的转录组学分析揭示了 NOX4-/- 小鼠 PH 后的显着变化，并支持Myc在增殖相关基因调控节点中的相关作用。有趣的是，RNA-seq 还揭示了与 TGF-β 通路激活相关的基因表达的变化。事实上，在 NOX4 缺失小鼠中，活性 TGF-β1 的水平、Smads 的磷酸化及其靶 p21 的水平在 24 小时时较低。Nox4 似乎对于体内肝再生的终止不是必需的，对于体外肝细胞对 TGF-β1 的生长抑制反应也不是必需的，这表明它有可能作为改善肝再生的治疗靶点而没有副作用。