Periplaneta americana extract attenuates hepatic fibrosis progression by inhibiting collagen synthesis and regulating the TGF-β1/Smad signaling pathway

Introduction. Liver fibrosis is the damage repair response following chronic liver diseases. Activated hepatic stellate cells (HSCs) are the main extracellular matrix (ECM)-producing cells and key regulators in liver fibrosis. Periplaneta americana shows prominent antifibrotic effects in liver fibrosis; however, the underlying mechanisms remain undetermined. This study aimed to elucidate the therapeutic effects of P. americana extract (PA-B) on liver fibrosis based on the regulation of the TGF-β1/Smad signal pathway. Material and methods. HSCs and Sprague Dawley rats were treated with TGF-β1 and CCl4, respectively, to establish the hepatic fibrosis model in vitro and in vivo. The effect of PA-B on liver rat fibrosis was evaluated by biochemical (serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), hyaluronic acid (HA), laminin (LN), collagen type Ⅳ (Col-Ⅳ), pro-collagen type Ⅲ (PC-Ⅲ)) and histological examinations. Further, fibrogenic markers expression of alpha smooth muscle actin (α-SMA), collagen type I (Col-I), and collagen type III (Col-III), and the TGF-β1/Smad pathway-related factors were assessed by immunofluorescence (IF), real time quantitative polymerase chain reaction (RT-qPCR), and western blotting (WB). Results. Treatment of HSC-T6 cells with PA-B suppressed the expression of α-SMA, Col-I, and Col-III, downregulated the expression of TGF-β1 receptors I and II (TβR I and TβR II, respectively), Smad2, and Smad3, and upregulated Smad7 expression. PA-B mitigates pathologic changes in the rat model of liver fibrosis, thus alleviating liver index, and improving liver function and fibrosis indices. The effects of PA-B on the expression of α-SMA, Col-I, Col-III, TβR I, TβR II, Smad2, Smad3, and Smad7 were consistent with the in vitro results, including reduced TGF-β1 expression. Conclusions. The therapeutic effect of PA-B on liver fibrosis might involve suppression of the secretion and expression of TGF-β1, regulation of the TGF-β1/Smad signaling pathway, and inhibition of collagen production and secretion

Full-Length Dystrophin Restoration via Targeted Exon Addition in DMD-Patient Specific iPSCs and Cardiomyocytes

Duchenne muscular dystrophy (DMD) is the most common fatal muscle disease, with an estimated incidence of 1/3500–1/5000 male births, and it is associated with mutations in the X-linked DMD gene encoding dystrophin, the largest known human gene. There is currently no cure for DMD. The large size of the DMD gene hampers exogenous gene addition and delivery. The genetic correction of DMD patient-derived induced pluripotent stem cells (DMD-iPSCs) and differentiation into suitable cells for transplantation is a promising autologous therapeutic strategy for DMD. In this study, using CRISPR/Cas9, the full-length dystrophin coding sequence was reconstructed in an exon-50-deleted DMD-iPSCs by the targeted addition of exon 50 at the junction of exon 49 and intron 49 via homologous-directed recombination (HDR), with a high targeting efficiency of 5/15, and the genetically corrected iPSCs were differentiated into cardiomyocytes (iCMs). Importantly, the full-length dystrophin expression and membrane localization were restored in genetically corrected iPSCs and iCMs. Thus, this is the first study demonstrating that full-length dystrophin can be restored in iPSCs and iCMs via targeted exon addition, indicating potential clinical prospects for DMD gene therapy