Yuduo Gao,Yiqing Chen,Zhenghao Hu,Ziyue Wang,Dongchang Sun,Chengfan Wu,Qi Shuai,Yunfeng Yan
Heat shock protein 47 (HSP47) regulates collagen synthesis and fibrosis, which makes it a promising target for controlling pulmonary fibrosis. However, challenges such as cell-selective and efficient drug delivery continue to impede the verification and translation of HSP47-targeted therapies. The CRISPR/Cas9 system can effectively inhibit the overexpression of HSP47 within fibrotic lesions while significantly reducing off-target effects. This study develops a class of lipid nanoparticles (LNPs) capable of codelivering Cas9 mRNA and sgHSP47 to fibroblasts. By incorporation of an additional cationic effector molecule, 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP), these LNPs achieve differential delivery across various cell lines and markedly enhance uptake efficiency in fibroblasts. This system mediates effective HSP47 knockdown in L929 cells and the lungs of C57BL/6 mice, leading to reduced collagen deposition and fibroblast activation in bleomycin-induced idiopathic pulmonary fibrosis (IPF) mice. This LNP platform holds substantial potential for gene editing-based IPF therapy and provides valuable insights for the efficient and selective delivery of CRISPR/Cas9 systems.
Background Biglycan (BGN) is a small leucine-rich proteoglycan that participates in the production of excess extracellular matrix (ECM) and is related to fibrosis in many organs. However, the role of BGN in liver fibrosis remains poorly understood. This study aimed to investigate the role and mechanism of BGN in liver fibrosis. Methods Human liver samples, Bgn −/0 (BGN KO) mice and a human LX-2 hepatic stellate cells (HSCs) model were applied for the study of experimental fibrosis. GEO data and single-cell RNA-seq data of human liver tissue were analysed as a bioinformatic approach. Coimmunoprecipitation, immunofluorescence staining, western blotting and qRT–PCR were conducted to identify the regulatory effects of BGN on heat shock protein 47 (HSP47) expression and liver fibrosis. Results We observed that hepatic BGN expression was significantly increased in patients with fibrosis and in a mouse model of liver fibrosis. Genetic deletion of BGN disrupted TGF-β1 pathway signalling and alleviated liver fibrosis in mice administered carbon tetrachloride (CCl 4 ). siRNA-mediated knockdown of BGN significantly reduced TGF-β1-induced ECM deposition and fibroblastic activation in LX-2 cells. Mechanistically, BGN directly interacted with and positively regulated the collagen synthesis chaperon protein HSP47. Rescue experiments showed that BGN promoted hepatic fibrosis by regulating ECM deposition and HSC activation by positively regulating HSP47. Conclusion Our data indicate that BGN promotes hepatic fibrosis by regulating ECM deposition and HSC activation through an HSP47-dependent mechanism. BGN may be a new biomarker of hepatic fibrosis and a novel target for disease prevention and treatment.
Intestinal fibrosis is a common complication of inflammatory bowel disease. There is still a lack of effective drugs for the prevention or treatment of intestinal fibrosis. Heat shock protein 47 (HSP47) plays a key role in the development of intestinal fibrosis. In this study we investigated the therapeutic potential and underlying mechanisms of fraxinellone, a degraded limonoid isolated from the root bark of Dictamnus dasycarpus , in the treatment of intestinal fibrosis. Intestinal fibrosis was induced in mice by dextran sodium sulfate (DSS) treatment. DDS-treated mice were administered fraxinellone (7.5, 15, 30 mg·kg −1 ·d −1 , i.g .) for 45 days. We showed that fraxinellone administration dose-dependently alleviated DSS-induced intestinal impairments, and reduced the production of intestinal fibrosis biomarkers such as α-smooth muscle actin (SMA), collagen I, hydroxyproline, fibronectin and laminin, and cytokines such as TGF-β, TNF-α and IL-β. We then established in vitro intestinal fibrosis cell models in SW480 and HT-29 cells, and demonstrated that treatment with fraxinellone (3, 10, 30 μM) significantly relieved TGF-β-induced fibrosis responses by inhibiting the TGF-β/Smad2/3 signaling pathway. Molecular docking suggested that the fraxinellone might disrupt the interaction between HSP47 and collagen, which was confirmed by coimmunoprecipitation experiments. SPR analysis showed that fraxinellone had a high affinity for HSP47 with a K d value of 3.542 × 10 −5 M. This study provides a new example of HSP47-collagen intervention by a natural compound and has important implications for the clinical treatment of inflammation-induced issue fibrosis.
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