Abstract

Background & Aims: Liver fibrosis is a critical pathological process in the progression of chronic liver diseases into severe forms, liver cirrhosis, liver failure, and liver cancer, posing a major global health burden with approximately 2 million deaths each year. Yet its incomplete mechanistic understanding hinders the development of effective therapies. Stress response signaling has been found to play a role in fibrosis development. Traditional and complementary medicine have long been widely explored for anti-fibrotic treatments. Huaji Ruangan Decoction (HJRGD), a classic traditional Chinese medicinal formula, has shown therapeutic promise in treating liver fibrosis. However, its mechanisms of action remain largely unknown.

Methods: We used a mouse model of liver fibrosis induced by intraperitoneal injection of carbon tetrachloride (CCl4) to evaluate the effect of HJRGD on liver injury, inflammation, and fibrosis. Additionally, we used tissue-specific stress response protein ATF4-deficient mice to examine the role of stress signaling in fibrosis development. Histopathological analyses, including H&E, Sirius Red, Masson, immunohistochemical staining, were conducted on mice livers to evaluate the therapeutic effects of HJRGD. Western blot, qPCR analysis, and biochemical analyses of sera and livers to explore the mechanisms underlying the anti-fibrotic effects of HJRGD. Primary hepatic stellate cells (HSCs) were isolated and cultured to further elucidate the mechanisms. Moreover, UHPLC-MS and computer-aided drug analysis were utilized to identify the potential active components of HJRGD contributing to its therapeutic effects on liver fibrosis.

Results: HJRGD markedly decreased collagen and extracellular matrix deposition by inhibiting hepatic stellate cell activation in CCl4-induced mice livers, accompanied with alleviated liver pathological changes including liver injury and inflammation. ATF4 played an critical role in the anti-fibrotic effect of HJRGD, as ATF4-deficient mice exhibited worsened liver inflammation and fibrosis, and notably, the beneficial effects of HJRGD were abolished in these mice. Furthermore, a comprehensive analysis of the active components of HJRGD strongly indicates that some bioactive ingredients of HJRGD modulate ATF4 signaling to mitigate liver fibrosis.

Conclusion: Our data demonstrate a robust protective anti-fibrosis effect of HJRGD through the regulation of ATF4-mediated stress signaling in HSCs and hepatocytes. Notably, certain active ingredients of HJRGD show potential as effective modulators of ATF4 stress response signaling pathway to reverse the fibrosis, meriting further investigation in the future. These results signify a novel approach to anti-fibrosis treatment and shed the light for developing new therapeutic strategies.

Abstract

Demyelination is the hallmark of multiple sclerosis (MS). Promoting remyelination is an important strategy to treat MS. Astragalus polysaccharides (APS) is the main bioactive component of Astragalus membranaceus, which is medical herb used for treating neurological disorders based on the clinical experiences in traditional Chinese medicine. To investigate the effects of APS on demyelination/remyelination and the underlying mechanisms, Experimental autoimmune encephalomyelitis (EAE) and cuprizone-induced demyelination model in mice were set up and treated with APS. It was found that APS could prevent demyelination in EAE mice, could relieve the neurobehavioral dysfunctions caused by demyelination, and efficaciously could facilitate remyelination in vivo. In order to determine whether the mechanism of enhancing remyelination was associated with the differentiation of neural stem cells (NSCs), biomarkers of NSCs, astrocytes, oligodendrocytes and neurons were measured in the corpus callosum tissues of mice through Real-time PCR, Western blot and immunohistochemistry assays. Data revealed that APS suppressed the stemness of NSCs, reduced the differentiation of NSCs into astrocytes, and promoted the differentiation into oligodendrocytes and neurons. This phenomenon was confirmed in the differentiation model of C17.2 NSCs cultured in vitro. Since Sonic hedgehog signaling pathway has been proven to be crucial to the differentiation of NSCs into oligodendrocytes, we examined expression levels of the key molecules in this pathway in vivo and in vitro, and eventually found APS activated this signaling pathway. Together, our results demonstrated that APS probably activated Sonic hedgehog signaling pathway first, then induced NSCs to differentiate into oligodendrocytes and promoted remyelination, which suggested that APS might be a potential candidate in treating autoimmune inflammatory diseases in the central nervous system (CNS) such as MS.

Abstract

The growing demand for natural anti-aging ingredients necessitates scientific validation of traditional cosmetic materials. Multani Mitti (MM), a clay widely used in South Asian traditional skincare, lacks comprehensive chemical and biological characterization. This study employed a multi-analytical approach to investigate MM’s anti-aging potential through chemical analysis, enzyme inhibition studies, and in silico evaluations. Five commercial MM samples were pooled and analyzed using instrumental neutron activation analysis (INAA) and Gas Chromatography–Mass Spectrometry (GC-MS). INAA revealed silicon as the predominant inorganic constituent (169.3742 mg/g), while GC-MS identified 13 bioactive compounds, with Beta-sitosterol (15.45% area), Docosanamide (12.36% area), and Cyclohexasiloxane (9.80% area) being the most abundant. MM demonstrated significant enzyme inhibition against key aging-related enzymes, with notably strong effects on hyaluronidase (IC50: 18 μg/mL) and tyrosinase (IC50: 27 μg/mL), outperforming standard inhibitors. The antioxidant activity showed moderate effectiveness (IC50: 31.938 μg/mL) compared to ascorbic acid (IC50: 8.5 μg/mL). Molecular docking studies of identified compounds against hyaluronidase (PDB: 1FCV) and tyrosinase (PDB: 3NQ1) revealed Beta-sitosterol and Benzyl-piperazine-carboxamide as the most promising candidates, showing strong binding affinities (−8.5 and −8.6 kcal/mol, respectively) and favorable ADMET profiles. This comprehensive characterization provides the first scientific evidence supporting MM’s traditional use in skincare and identifies specific compounds that may contribute to its anti-aging properties, warranting further investigation for modern cosmetic applications.

Abstract

Dementia, particularly Alzheimer’s disease, is a progressive neurodegenerative disorder characterized by cognitive decline and limited treatment options due to adverse effects of current drugs. Majoon Boolis, a traditional Unani polyherbal formulation comprising thirteen medicinal herbs, has historically been used as a neuroprotective agent, yet its therapeutic relevance in dementia remains scientifically unverified. This study employed a comprehensive computational approach integrating network pharmacology, molecular docking, and molecular dynamics (MD) simulations to investigate the neuroprotective potential of Majoon Boolis. A total of 666 phytoconstituents were identified through literature mining and retrieved from ChemSpider databases. Target prediction and gene mapping revealed dementia-associated genes, and protein–protein interaction analysis identified 28 hub genes, including Casp3, Il6, Akt1, and Mapk3. Gene ontology and KEGG pathway enrichment analysis showed significant involvement in oxidative stress regulation, apoptosis, and neuroinflammatory processes, with the TNF signalling pathway emerging as a crucial pathway. Caspase-3 was selected as the key therapeutic target based on its central role in apoptosis. Molecular docking identified jeediflavanone and cuscutoside B as top binders to caspase-3, with cuscutoside B demonstrating favourable molecular interactions with the receptor. Further, 100-nanosecond MD simulations confirmed that cuscutoside B maintained stable binding and favourable conformational dynamics within the active site, comparable to the co-crystallized ligand B92. These results highlight the potential of Majoon Boolis, particularly cuscutoside B, as a promising natural candidate for dementia therapy. The study emphasizes the value of integrating traditional medicinal knowledge with modern computational tools for identifying novel neuroprotective agents and guiding future experimental and clinical investigations.