Excessive production of transforming growth factor β1 (TGF-β1) in activated hepatic stellate cells (aHSCs) promotes liver fibrosis by activating the TGF-β1/Smad signaling pathway. Thus, specifically inhibiting the pro-fibrotic activity of TGF-β1 in aHSCs is an ideal strategy for treating liver fibrosis. Overexpression of platelet-derived growth factor β receptor (PDGFβR) has been demonstrated on the surface of aHSCs relative to normal cells in liver fibrosis. Interferon-gamma peptidomimetic (mIFNγ) and truncated TGF-β receptor type II (tTβRII) inhibit the TGF-β1/Smad signaling pathway by different mechanisms. In this study, we designed a chimeric protein by the conjugation of (1) mIFNγ and tTβRII coupled via plasma protease-cleavable linker sequences (FNPKTP) to (2) PDGFβR-recognizing peptide (BiPPB), namely BiPPB-mIFNγ-tTβRII. This novel protein BiPPB-mIFNγ-tTβRII was effectively prepared using Escherichia coli expression system. The active components BiPPB-mIFNγ and tTβRII were slowly released from BiPPB-mIFNγ-tTβRII by hydrolysis using the plasma protease thrombin in vitro. Moreover, BiPPB-mIFNγ-tTβRII highly targeted to fibrotic liver tissues, markedly ameliorated liver morphology and fibrotic responses in chronic liver fibrosis mice by both inhibiting the phosphorylation of Smad2/3 and inducing the expression of Smad7. Meanwhile, BiPPB-mIFNγ-tTβRII markedly reduced the deposition of collagen fibrils and expression of fibrosis-related proteins in acute liver fibrosis mice. Furthermore, BiPPB-mIFNγ-tTβRII showed a good safety performance in both liver fibrosis mice. Taken together, BiPPB-mIFNγ-tTβRII improved the in vivo anti-liver fibrotic activity due to its high fibrotic liver-targeting potential and the dual inhibition of the TGF-β1/Smad signaling pathway, which may be a potential candidate for targeting therapy on liver fibrosis.

活化的肝星状细胞 (aHSC) 中过量产生转化生长因子 β1 (TGF-β1)，通过激活 TGF-β1/Smad 信号通路促进肝纤维化。因此，特异性抑制aHSCs中TGF-β1的促纤维化活性是治疗肝纤维化的理想策略。与肝纤维化中的正常细胞相比，aHSC 表面存在血小板衍生生长因子 β 受体 (PDGFβR) 的过度表达。干扰素-γ拟肽 (mIFNγ) 和截短型 TGF-β 受体 II (tTβRII) 通过不同机制抑制 TGF-β1/Smad 信号通路。在本研究中，我们设计了一种嵌合蛋白，将 (1) mIFNγ 和 tTβRII 通过血浆蛋白酶可裂解接头序列 (FNPKTP) 偶联至 (2) PDGFβR 识别肽 (BiPPB)，即 BiPPB-mIFNγ-tTβRII。这种新型蛋白BiPPB-mIFNγ-tTβRII是利用大肠杆菌表达系统有效制备的。体外血浆蛋白酶凝血酶水解后，BiPPB-mIFNγ-tTβRII 缓慢释放活性成分 BiPPB-mIFNγ 和 tTβRII。此外，BiPPB-mIFNγ-tTβRII高度靶向纤维化肝组织，通过抑制Smad2/3的磷酸化和诱导Smad7的表达，显着改善慢性肝纤维化小鼠的肝脏形态和纤维化反应。同时，BiPPB-mIFNγ-tTβRII显着减少急性肝纤维化小鼠中胶原纤维的沉积和纤维化相关蛋白的表达。此外，BiPPB-mIFNγ-tTβRII 在两只肝纤维化小鼠中均表现出良好的安全性。综上所述，BiPPB-mIFNγ-tTβRII 由于其高纤维化肝靶向潜力和对 TGF-β1/Smad 信号通路的双重抑制，提高了体内抗肝纤维化活性，可能是靶向治疗的潜在候选者。肝纤维化。