Figures and original data

Liver fibrosis is a dynamic and potentially reversible pathological consequence of chronic liver injury, in which persistent oxidative stress and inflammatory cascades drive progressive extracellular matrix deposition. Cyclic nitroxide radicals have diverse biological activities in vitro and in vivo; however, the effects of these stable radicals on liver fibrosis and other liver inflammatory diseases remain unclear. This study systematically evaluates 3-carbamoyl proxyl nitroxide (3-CP), a stable nitroxide compound, for its therapeutic potential against carbon tetrachloride (CCl₄)-induced liver fibrosis. In vitro, 3-CP inhibited the activation, migration, and proliferation of hepatic stellate cells (HSCs), and suppressed the expression of α-smooth muscle actin (α-SMA) and collagen I (COL1). In a BALB/c mouse model of CCl₄-induced liver fibrosis, 20 and 40 mg/kg 3-CP reduced the fibrosis area from 13.6 ± 1.0% (model group) to 6.9 ± 0.9% and 5.7 ± 1.3%, respectively. This was corroborated by decreased serum alanine and aspartate aminotransferase levels, restored liver architecture, and diminished collagen deposition. Mechanistically, 3-CP modulated the TLR4/NF-κB signaling pathway through the downregulation of phosphorylated NF-κB p65 (p-p65), which aligns with the results of molecular docking analyses. The relative mRNA levels of interleu-1β (IL-1β), interleu-6 (IL-6), tumor necrosis factor-α (TNF-α) and transforming growth factor-β (TGF-β) were decreased by 0.99 ± 0.04%, 1.01 ± 0.03%, 0.97 ± 0.04%, and 1.02 ± 0.06%, respectively. These findings indicated that 3-CP is a therapeutic agent that concurrently addresses oxidative damage and inflammatory signaling during fibrogenesis.

肝纤维化（Liver fibrosis）是慢性肝损伤所致的动态且具备可逆潜能的病理性转归，在此过程中持续性氧化应激与炎症级联反应会驱动细胞外基质进行性沉积。环状氮氧自由基（Cyclic nitroxide radicals）在体外与体内均具有多样的生物学活性，但这类稳定自由基对肝纤维化及其他肝脏炎症性疾病的作用仍不明确。本研究系统评估了3-氨甲酰基普罗克斯尔氮氧自由基（3-carbamoyl proxyl nitroxide，简称3-CP）——一种稳定的氮氧自由基化合物——对抗四氯化碳（carbon tetrachloride，CCl₄）诱导的肝纤维化的治疗潜力。体外实验中，3-CP可抑制肝星状细胞（hepatic stellate cells，HSCs）的活化、迁移与增殖，并下调α-平滑肌肌动蛋白（α-smooth muscle actin，α-SMA）与I型胶原（collagen I，COL1）的表达。在BALB/c小鼠CCl₄诱导的肝纤维化模型中，20 mg/kg与40 mg/kg剂量的3-CP可将纤维化面积从模型组的13.6±1.0%分别降至6.9±0.9%与5.7±1.3%。该结论通过血清丙氨酸氨基转移酶（alanine aminotransferase，ALT）与天冬氨酸氨基转移酶（aspartate aminotransferase，AST）水平下调、肝脏组织结构恢复以及胶原沉积减少得到了证实。从机制层面来看，3-CP可通过下调磷酸化NF-κB p65（phosphorylated NF-κB p65，简称p-p65）的表达调控TLR4/NF-κB信号通路，这一结果与分子对接分析的结论相符。白细胞介素-1β（IL-1β）、白细胞介素-6（IL-6）、肿瘤坏死因子-α（TNF-α）以及转化生长因子-β（TGF-β）的相对mRNA表达水平分别下降了0.99±0.04%、1.01±0.03%、0.97±0.04%与1.02±0.06%。上述研究结果表明，3-CP是一种可同时针对纤维化发生过程中氧化损伤与炎症信号通路发挥作用的治疗候选药物。