The present research focused on estimating, for the first time, the potential protective effects of bromelain against D-galactosamine-induced acute liver injury in rats as well as identifying the possible underlying mechanisms. Silymarin (100 mg/kg/day, p.o.) as a reference drug or bromelain (20 and 40 mg/kg/day, p.o.) were administered for 10 days, and on the 8th day of the experiment, a single dose of galactosamine (400 mg/kg/i.p.) induced acute liver injury. Pretreatment with bromelain improved liver functions and histopathological alterations induced by galactosamine. Bromelain ameliorated oxidative stress by inducing SIRT1 protein expression and increasing LKB1 content. This resulted in phosphorylating the AMPK/GSK3β axis, which stimulated Nrf2 activation in hepatic cells and thus increased the activity of its downstream antioxidant enzymes [HO-1 and NQO1]. Besides, bromelain exerted significant anti-apoptotic and anti-inflammatory effects by suppressing hepatic contents of TNF-α, NF-κB p65, as well as caspase-8 and caspase-9. The protective effects of bromelain₄₀ were proved to be better than silymarin and bromelain₂₀ in most of the assessed parameters. Our results highlight the significant hepatoprotective effects of bromelain against acute liver injury through modulation of SIRT1/LKB1/AMPK, GSK3β/Nrf2 signalling in addition to NF-κB p65/TNF-α/ caspase-8 and -9 pathway.

Gastrointestinal (GI) diseases have become a global health issue and an economic burden due to their wide distribution, late prognosis, and the inefficacy of recent available medications. Therefore, it is crucial to search for new strategies for their management. In the recent decades, mesenchymal stem cells (MSCs) therapy has attracted attention as a viable option for treating a myriad of GI disorders such as hepatic fibrosis (HF), ulcerative colitis (UC), acute liver injury (ALI), and non-alcoholic fatty liver disease (NAFLD) due to their regenerative and paracrine properties. Importantly, recent studies have shown that MSC-derived extracellular vesicles (MSC-EVs) are responsible for most of the therapeutic effects of MSCs. In addition, EVs have revealed several benefits over their parent MSCs, such as being less immunogenic, having a lower risk of tumour formation, being able to cross biological barriers, and being easier to store. MSC-EVs exhibited regenerative, anti-oxidant, anti-inflammatory, anti-apoptotic, and anti-fibrotic effects in different experimental models of GI diseases. However, a key issue with their clinical application is the maintenance of their stability and efficacy following in vivo transplantation. Preconditioning of MSC-EVs or their parent cells is one of the novel methods used to improve their effectiveness and stability. Herein, we discuss the application of MSC-EVs in several GI disorders taking into account their mechanism of action. We also summarise the challenges and restrictions that need to be overcome to promote their clinical application in the treatment of various GI diseases as well as the recent developments to improve their effectiveness. Graphical abstract A representation of the innovative preconditioning techniques that have been suggested for improving the therapeutic efficacy of MSC-EVs in GI diseases. The pathological conditions in various GI disorders (ALI, UC, HF and NAFLD) create a harsh environment for EVs and their parents, increasing the risk of apoptosis and senescence of MSCs and thereby diminishing MSC-EVs yield and restricting their large-scale applications. Preconditioning with pharmacological agents or biological mediators can improve the therapeutic efficacy of MSC-EVs through their adaption to the lethal environment to which they are subjected. This can result in establishment of a more conducive environment and activation of numerous vital trajectories that act to improve the immunomodulatory, reparative and regenerative activities of the derived EVs, as a part of MSCs paracrine system. ALI, acute liver injury; GI diseases, gastrointestinal diseases; HF, hepatic fibrosis; HSP, heat shock protein; miRNA, microRNA; mRNA, messenger RNA; MSC-EVs, mesenchymal stem cell-derived extracellular vesicles; NAFLD, non-alcoholic fatty liver disease; UC, ulcerative colitis.

Hepatic fibrosis is one of the major worldwide health concerns which requires tremendous research due to the limited outcomes of the current therapies. The present study was designed to assess, for the first time, the potential therapeutic effect of rupatadine (RUP) in diethylnitrosamine (DEN)-induced liver fibrosis and to explore its possible mechanistic actions. For the induction of hepatic fibrosis, rats were treated with DEN (100 mg/kg, i.p.) once weekly for 6 consecutive weeks, and on the 6th week, RUP (4 mg/kg/day, p.o.) was administered for 4 weeks. Treatment with RUP ameliorated changes in body weights, liver indices, liver function enzymes, and histopathological alterations induced by DEN. Besides, RUP amended oxidative stress, which led to the inhibition of PAF/NF-κB p65-induced inflammation, and, subsequently, prevention of TGF-β1 elevation and HSCs activation as indicated by reduced α-SMA expression and collagen deposition. Moreover, RUP exerted significant anti-fibrotic and anti-angiogenic effects by suppressing Hh and HIF-1α/VEGF signaling pathways. Our results highlight, for the first time, a promising anti-fibrotic potential of RUP in rat liver. The molecular mechanisms underlying this effect involve the attenuation of PAF/NF-κB p65/TGF-β1 and Hh pathways and, subsequently, the pathological angiogenesis (HIF-1α/VEGF). Graphical abstract

Liver fibrosis is still a serious health concern worldwide, and there is increasing interest in mesenchymal stem cells (MSCs) with tremendous potential for treating this disease because of their regenerative and paracrine effects. Recently, many researches have focused on using the released exosomes (EXOs) from stem cells to treat liver fibrosis rather than using parent stem cells themselves. MSC-derived EXOs (MSC-EXOs) have demonstrated favourable outcomes similar to cell treatment in terms of regenerative, immunomodulatory, anti-apoptotic, anti-oxidant, anti-necroptotic, anti-inflammatory and anti-fibrotic actions in several models of liver fibrosis. EXOs are superior to their parent cells in several terms, including lower immunogenicity and risk of tumour formation. However, maintaining the stability and efficacy of EXOs after in vivo transplantation remains a major challenge in their clinical applicability. Therefore, several strategies have been applied in EXOs engineering, such as parental cell modification or modifying EXOs directly to achieve optimum performance of EXOs in treating liver fibrosis. Herein, we discuss the underlying mechanisms of liver fibrosis with an overview of the available therapies, among them EXOs. We also summarise the recent developments in improving the effectiveness of EXOs with the advantages and limitations of these approaches in terms of the upcoming clinical applications. Graphical Abstract

Background: The cardiotoxic effects of doxorubicin (DOX), a powerful chemotherapeutic drug, are widely recognized. Cuproptosis, a unique copper-dependent form of controlled cell death, may be involved in DOX-induced cardiomyopathy, according to recent findings. This study employs both in vivo and in silico procedures to investigate the protective effects of the copper chelator penicillamine (PEN) and the role of cuproptosis in DOX-related cardiotoxicity. Methods: Thirty-two adult Sprague Dawley rats were allocated into four groups (n = 8): control, DOX, DOX+PEN, and PEN. Cardiac function was assessed via echocardiography. Serum cardiac biomarkers (LDH, CK-MB, CTnI), oxidative stress markers (SOD, GPX, MDA), and expression levels of cuproptosis-related genes (FDX1, LIAS, SLC31A1, ATP7A) were evaluated. Histopathological examinations and immunohistochemical staining for FDX1, SLC31A1, and DLAT were performed. Molecular docking simulated PEN’s interaction with cuproptosis-related proteins. Network pharmacology and molecular docking studies were also conducted to identify core molecular targets and simulate PEN’s binding interactions with key cuproptosis regulators. Results: DOX administration induced significant cardiac dysfunction, oxidative stress, and upregulation of cuproptosis markers. PEN treatment mitigated these effects, improved cardiac function, reduced fibrosis, and suppressed the expression of cuproptosis-related genes and proteins. Docking results confirmed strong interactions between PEN and cuproptosis-regulatory proteins. Network pharmacology revealed 14 key overlapping targets linking PEN with cuproptosis and DOX-induced cardiotoxicity. Conclusion: This study provides experimental evidence implicating cuproptosis in DOX-induced cardiomyopathy. PEN exerts cardioprotection, potentially by targeting this pathway, offering a promising therapeutic strategy.