Background: Indole phytoalexins, plant-derived compounds present in cruciferous vegetables, have demonstrated anticancer properties. Brassinin (BSN), derived from Brassica campestris L. var. campestris, is known for its potent antitumor effects on various cancers. However, the role of ferroptosis in regulating the antitumor effects of BSN has not been fully elucidated.

Methods: The components of B. campestris L. against colorectal cancer (CRC) were analyzed by network pharmacology. CCK-8 assay and colony formation assay detected cell viability induced by BSN. Molecular docking verified the binding of BSN to the target protein. Western blot and reverse transcription–quantitative polymerase chain reaction (RT-qPCR) assay revealed whether BSN can inactivate the NRF2 signaling and inhibit the expression of p62 and HO-1. The RKO-xenograft tumor models were established and then were treated by 75 or 150 mg/kg BSN to verify the antitumor efficacy and side effects of BSN.

Results: Network pharmacology suggested that BSN is the most important component of B. campestris L. against CRC. BSN inhibits CRC cell viability in a dose- and time-dependent manner. Furthermore, this inhibitory effect is associated with the induction of ferroptosis, as BSN suppresses the cell viability of CRC by inducing GPX4-regulated ferroptosis. BSN may bind to NRF2 protein to inactivate the NRF2 signaling, inhibiting the expression of p62 and HO-1. Importantly, a low dose or a high dose of BSN significantly reduced the tumor growth in vivo.

Conclusions: Our findings reveal that BSN blocks CRC growth by inducing p62/NRF2/GPX4-regulated ferroptosis, which may be a novel lead compound for tumor treatment.

Background: Colocasia esculenta (L.) Schott, known as the taro vegetable, possesses various beneficial effects and is traditionally used in folk medicine. This study explores the ameliorative antioxidant and hepatoprotective effect of a methanolic extract of the C. esculenta flower (ME-CEF) against oxidative damage and hepatotoxicity in mice.

Methods: The antioxidant efficacy of ME-CEF was assessed using 2,2′-azino-bis-(3-ethylbenzothiazoline-6-sulfonic) (ABTS) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging assay. The hepatoprotective effect was investigated by an assessment of liver injury indicators (amino transferase [ALT], aspartate amino transferase [AST], alkaline phosphatase [ALP], bilirubin, creatinine) and normalizing lipid profiles (cholesterol [CHO], triglyceride [TG], high-density lipoprotein [HDL], and low-density lipoprotein [LDL]) along with histopathological study and antioxidant enzymes (CAT). A phytochemical analysis, both qualitative and quantitative, was conducted, including gas chromatography-tandem mass spectrometry (GC–MS/MS) analysis and an in silico molecular docking study.

Results: The Result Showed that ME-CEF Possesses Moderate ABTS and DPPH Scavenging Activity with IC₅₀ Values of 117.18 and 160.41 μg/mL. As Illustrated by Reducing Liver Enzymes (ALT, AST, ALP, Bilirubin, Creatinine) and Lipid Profile (CHO, TG, LDL) and Raising HDL Levels (p < 0.01), ME-CEF Dose Dependently Mitigated CCl₄-Induced Acute Liver Injury. Furthermore, ME-CEF Blocked Hepatic Oxidative Stress by Boosting Antioxidant Enzymes (CAT) and Preventing Liver Tissue Damage and Apoptosis. In Silico Investigations Also Showed a Promising Binding Affinity with Tumor Necrosis Factor α (TNF-α), Interleukin 6 (IL-6), PRAP-1, and Xanthin Oxidoreductase, which Displayed Antioxidant and Hepatoprotective Candidacy while Notable Safety and Efficacy Profile Was Also Documented through ADME/T Studies. Histopathological Analysis Showed Reduced Hepatocellular Necrosis and Vascular Congestion in Silymarin and Extract Groups.

Conclusion: Based on these results, our findings strongly recommend the medicinal use of the plant, highlighting its antioxidant and hepatoprotective potentials.

Background: Silicosis is an occupational lung disease that is caused by chronic exposure to silica dust. Silica-exposed workers are at higher risk of developing TB, resulting in lung fibrosis and significant respiratory dysfunction. Diosgenin is a steroidal saponin that has been shown to exert a therapeutic effect on lung injury. Therefore, we investigated the potential efficacy of diosgenin in treating silicotuberculosis by evaluating its effectiveness against Mycobacterium smegmatis, as well as its antifibrotic and antioxidant effects in silica-induced TB in rats.

Methods: Silicosis was induced by intratracheal instillation of 50 mg/kg crystalline silica in Sprague–Dawley rats. Rats were grouped into 7 (10 per group). Different doses of diosgenin (1, 10, and 20 mg/kg) and saline were administered for 30 days. Afterwards, five rats from each group were sacrificed, and the five remaining rats in each group, except the control, received Mycobacterium smegmatis. Treatment continued until the 50th day, and the animals were sacrificed at the end of the experiment. The result was analyzed using a one-way analysis of variance (ANOVA) with GraphPad Prism.

Results: At a half-maximal inhibition concentration of 0.006043 μg/mL, diosgenin inhibited the growth of Mycobacterium smegmatis. Oxidative stress markers such as malondialdehyde were significantly reduced. The health-enhancing effects of catalase and superoxide dismutase were elevated. Additionally, histological findings demonstrated a significant improvement in respiratory function following diosgenin treatment.

Conclusion: Diosgenin treatment inhibited the growth of Mycobacterium smegmatis, leading to a reduction in the susceptibility of rats to infection and improved pulmonary function through its antioxidant effect.

Background: There are many forms of anabolic steroids, including stanozolol (Winstrol), which are popular for their muscle-building effects but dangerous to the heart. This present work is aimed at evaluating the pharmacologica impact of allicin, a natural attribute obtained from garlic, on obstructing cardiac injury in rabbits that received stanozolol.

Methods: Thirty rabbits were divided into three groups: control, stanozolol-treated, and stanozolol plus allicin. Cardiac function was assessed by measuring troponin, creatine kinase (CK), Galectin-3, and GDF-15. Oxidative stress and antioxidant markers, including malondialdehyde (MDA), glutathione, and catalase, were analyzed. Inflammatory mediators such as C-reactive protein (CRP), interleukin-6 (IL-6), NF-κB, iNOS, nitric oxide (NO), tumor necrosis factor-alpha (TNF-α), and interleukin-1 beta (IL-1β) were evaluated. Lipid profile parameters, including total cholesterol, low-density lipoprotein (LDL), and high-density lipoprotein (HDL), were measured. Histopathological examination assessed myocardial damage, fibrosis, and collagen deposition.

Results: Stanozolol administration significantly increased cardiac damage markers, oxidative stress, and inflammatory mediators while causing dyslipidemia, characterized by elevated LDL and total cholesterol and reduced HDL. Allicin co-administration effectively countered these effects by reducing oxidative stress and inflammation, restoring antioxidant balance, and improving lipid profiles. Histopathological analysis revealed severe myocardial disorganization, necrosis, and fibrosis in the stanozolol group, whereas the allicin-treated group exhibited preserved myocardial structure with reduced collagen deposition.

Conclusion: Allicin significantly mitigates stanozolol-induced cardiotoxicity by reducing oxidative stress, inflammation, lipid dysregulation, and myocardial damage, as evidenced by biochemical and histopathological findings. These results suggest that allicin may serve as a potential therapeutic agent to counteract the cardiovascular risks associated with anabolic steroid use.

Cisplatin chemotherapy has been used as the main treatment for different types of cancer. However, cisplatin chemotherapy-induced peripheral neuropathic pain (CIPNP) seriously affects the treatment process and quality of life of patients. In addition, it impacts the underlying mechanism and prevention and treatment strategies, indicating that drug selection and efficacy evaluation need to be further investigated. Furthermore, an animal model that is more consistent with the pathological mechanism needs to be developed. In this study, we describe and discuss the methods of developing and detecting CIPNP models in rats and mice induced by cisplatin chemotherapy. The aim was to improve the modeling rate and develop animal models that are more consistent with the developmental pattern of the disease. In addition, the study provides ideal reference animal models for clinical research and drug discovery and development.

Liver fibrosis, a hallmark pathological endpoint of chronic aging-related liver diseases, remains a clinical challenge with limited therapeutic options. In healthy liver, myeloid cells constitute <5% of total hepatic immune cells, primarily comprising tissue-resident Kupffer cells. However, during aging or chronic injury, bone marrow–derived myeloid cell recruitment increases by two- to threefold in murine fibrotic models, reaching 15%–20% of intrahepatic immune populations. These infiltrating myeloid subsets exhibit functional plasticity, dynamically differentiating into pro-inflammatory macrophages or fibrosis-promoting Kupffer-like cells, contingent upon chemokine gradients (e.g., CCL2/CCR2 axis) and damage-associated molecular patterns (DAMPs). This review systematically examines the regulatory mechanisms of myeloid cells in liver fibrogenesis, with particular emphasis on their developmental origins, hepatic recruitment dynamics, functional heterogeneity, and pathogenic contributions to fibrosis. Furthermore, signaling pathways involving myeloid cells in liver fibrosis and therapeutic approaches modulating their differentiation and recruitment are discussed in this review.

Lung cancer has one of the highest rates of incidence and mortality worldwide, making research on its mechanisms and treatments crucial. Animal models are essential in lung cancer research as they accurately replicate the biological characteristics and treatment outcomes seen in human diseases. Currently, various lung cancer models have been established, including chemical induction models, orthotopic transplantation models, ectopic transplantation models, metastasis models, and gene editing mouse models. Additionally, lung cancer grafts can be categorized into two types: tissue-based and cell-based grafts. This paper summarizes the phenotypes, advantages, and disadvantages of various induction methods based on their modeling techniques. The goal is to enhance the simulation of clinical lung cancer characteristics and to establish a solid foundation for future clinical research.

Background: Osteoarthritis (OA) is a common degenerative joint disease characterized by the progressive degradation of articular cartilage. Mitochondrial dysfunction and autophagy, including mitophagy, have been implicated in OA pathogenesis. Long noncoding RNAs (lncRNA) are emerging as key regulators in various cellular processes, but their roles in OA, particularly in chondrocytes, remain poorly understood. This study explores the involvement of lncRNA-GCH1 in regulating mitophagy and its impact on chondrocyte function and cartilage degradation in OA.

Methods: Primary chondrocytes were isolated from the cartilage tissues of OA patients and healthy controls. lncRNA-GCH1 expression was assessed using RNA-seq, reverse transcription quantitative polymerase chain reaction, and RNA fluorescence in situ hybridization. Functional assays, including Cell Counting Kit-8 (CCK-8), colony formation, flow cytometry, and Western blotting, were used to evaluate the effects of lncRNA-GCH1 knockdown on chondrocyte proliferation, apoptosis, cell cycle, and mitophagy. Mitochondrial function was assessed by measuring adenosine triphosphate production, reactive oxygen species levels, and mitochondrial membrane potential. In vivo, a murine OA model was used to examine the impact of lncRNA-GCH1 knockdown on cartilage degradation.

Results: lncRNA-GCH1 was upregulated in OA chondrocytes and localized in the cytoplasm. Knockdown of lncRNA-GCH1 enhanced cell proliferation and arrested cell cycle in G0/G1. It also suppressed mitophagy, improved mitochondrial function, and reduced matrix-degrading enzyme expression—effects that were reversed by rapamycin treatment. Meanwhile, lncRNA-GCH1 knockdown reduced PTEN-induced kinase 1 (PINK1) aggregation and in vivo local inhibition of PINK1 diminished cartilage degradation.

Conclusion: lncRNA-GCH1 regulates mitophagy in OA chondrocytes, influencing mitochondrial function and matrix degradation. Targeting lncRNA-GCH1 may offer a potential therapeutic approach for OA treatment.

Background: Metabolic abnormalities are considered to play a key regulatory role in vascular remodeling of pulmonary arterial hypertension. However, to date, there is a paucity of research documenting the changes in metabolome profiles within the supernatants of pulmonary artery smooth muscle cells (PASMC) during their transition from a contractile to a synthetic phenotype.

Methods: CCK-8 and Edu staining assays were used to evaluate the cell viability and proliferation of human PASMCs. IncuCyte ZOOM imaging system was used to continuously and automatically detect the migration of the PASMCs. A targeted metabolomics profiling was performed to quantitatively analyze 121 metabolites in the supernatant. Orthogonal partial least squares discriminant analysis was used to discriminate between PDGF-BB-induced PASMCs and controls. Metabolite set enrichment analysis was adapted to exploit the most disturbed metabolic pathways.

Results: Human PASMCs exhibited a transformation from contractile phenotype to synthetic phenotype after PDGF-BB induction, along with a significant increase in cell viability, proliferation, and migration. Metabolites in the supernatants of PASMCs treated with or without PDGF-BB were well profiled. Eleven metabolites were found to be significantly upregulated, whereas seven metabolites were downregulated in the supernatants of PASMCs induced by PDGF-BB compared to the vehicle-treated cells. Fourteen pathways were involved, and pyruvate metabolism pathway was ranked first with the highest enrichment impact followed by glycolysis/gluconeogenesis and pyrimidine metabolism.

Conclusions: Significant and extensive metabolic abnormalities occurred during the phenotypic transformation of PASMCs. Disturbance of pyruvate metabolism pathway might contribute to pulmonary vascular remodeling.

Background: This study investigated the impacts and mechanisms of yunweiling in the management of Functional Constipation (FC) using network pharmacology and experimental research.

Methods: Using the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP), Genecard, and Online Mendelian Inheritance in Man (OMIM) databases, a potential gene target for yunweiling in treating FC was found. A pharmacological network was built and viewed in Cytoscape. A protein interaction map was created with STRING and Cytoscape. ‘clusterProfiler’ helped uncover its mechanism. Molecular docking was done with AutoDock Vina. In a constipation mouse model, Western blot was used to assess yunweiling's effectiveness.

Results: To investigate yunweiling's therapeutic effects on FC, we employed a loperamide-induced constipation model. Successful model establishment was confirmed by first black stool time, reduced stool output, and impaired gastrointestinal motility. Yunweiling treatment, especially at high and medium doses, significantly alleviated constipation symptoms by reducing first black stool time, increasing stool output, and enhancing gastrointestinal motility. HE staining revealed yunweiling's ability to restore colon tissue structure. Yunweiling modulated the expression of key proteins TP53, P-AKT, P-PI3K, RET, and Rai, implicating its involvement in the PI3K-Akt signaling pathway. Comparative analysis showed yunweiling to be more effective than its individual components (shionone, β-sitosterol, and daucosterol) in improving constipation. The combination of yunweiling with TP53 and PI3K-Akt inhibitors further enhanced its therapeutic effects, suggesting a synergistic mechanism.

Conclusions: The integration of network pharmacology and experimental investigations indicated the effectiveness of yunweiling in managing FC, offering essential theoretical support for clinical application.

Background: Rabies virus (RABV)–derived neuronal tracing tools are extensively applied in retrograde tracing due to their strict retrograde transsynaptic transfer property and low neurotoxicity. However, the RABV infection and expression of fluorescence products would be gradually cleared while the infected neurons still survive, a phenomenon known as non-cytolytic immune clearance (NCLIC). This phenomenon introduced the risk of fluorescence loss and led to the omission of a subset of neurons that should be labeled, thereby interfering in the analysis of tracing results.

Methods: To compensate for the fluorescence loss problem, in this study, we developed a novel marker footprints (MF) mouse, involving a Cre recombinase-dependent red fluorescent reporter system and systemic expression of glycoprotein (G) and ASLV-A receptor (TVA). Using this mouse model combined with the well-developed RABV-EnvA-ΔG-GFP-Cre viral tool, we developed a novel green-to-red spectral labeling strategy.

Results: Neurons in the MF mouse could be co-labeled with green fluorescence from the very quick expression of the viral tool and with red fluorescence from the relatively slow expression of the neuron itself, so neurons undergoing NCLIC with green fluorescence loss could be relabeled red. Furthermore, newly infected neurons could be labeled green and other neurons could be labeled yellow due to the temporal expression difference between the two fluorescent proteins.

Conclusions: This is the first polysynaptic retrograde tracing labeling strategy that could label neurons using spectral fluorescence colors with only one injection of the viral tool, enabling its application in recognizing the labeling sequence of neurons in brain regions and enhancing the spatiotemporal resolution of neuronal tracing.

Background: New variants of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2) continue to drive global epidemics and pose significant health risks. The pathogenicity of these variants evolves under immune pressure and host factors. Understanding these changes is crucial for epidemic control and variant research.

Methods: Human angiotensin-converting enzyme 2(hACE2) transgenic mice were intranasally challenged with the original strain WH-09 and the variants Delta, Beta, and Omicron BA.1, while BALB/c mice were challenged with Omicron subvariants BA.5, BF.7, and XBB.1. To compare the pathogenicity differences among variants, we conducted a comprehensive analysis that included clinical symptom observation, measurement of viral loads in the trachea and lungs, evaluation of pulmonary pathology, analysis of immune cell infiltration, and quantification of cytokine levels.

Results: In hACE2 mice, the Beta variant caused significant weight loss, severe lung inflammation, increased inflammatory and chemotactic factor secretion, greater macrophage and neutrophil infiltration in the lungs, and higher viral loads with prolonged shedding duration. In contrast, BA.1 showed a significant reduction in pathogenicity. The BA.5, BF.7, and XBB.1 variants were less pathogenic than the WH-09, Beta, and Delta variants when infected in BALB/c mice. This was evidenced by reduced weight loss, diminished pulmonary pathology, decreased secretion of inflammatory factors and chemokines, reduced macrophage and neutrophil infiltration, as well as lower viral loads in both the trachea and lungs.

Conclusion: In hACE2 mice, the Omicron variant demonstrated the lowest pathogenicity, while the Beta variant exhibited the highest. Pathogenicity of the Delta variant was comparable to the original WH-09 strain. Among BALB/c mice, Omicron subvariants BA.5, BF.7, and XBB.1 showed no statistically significant differences in virulence.

Objective: Sheep are commonly used as large animal pre-clinical models for investigating cardiovascular therapies, interventions, anatomy and physiology. Further, novel small diameter vascular grafts are frequently tested via implantation into sheep carotid arteries (CAs). This is because, unlike humans, acute occlusion of one or both sheep CAs is not associated with morbidity or mortality and thus provides safer experimental testing, with reduced ethical constraints, animal numbers and costs. However, to date there has been no evidence regarding sheep tolerance of femoral artery (FA) occlusion.

Methods: In this study, seven sheep underwent CA graft surgery, with digital subtraction angiography (DSA) of the CAs performed every 2 months via femoral access, for a total of 8 months. Four months into the study, the left FA of two sheep became inaccessible due to a suspected FA occlusion. Thus, femoral angiography was performed, followed by FA dissection, FA histology and retrospective analysis of both veterinarian animal monitoring and pain scores.

Results: FA angiography and histology confirmed complete left FA occlusion in two sheep. Retrospective animal monitoring demonstrated sheep with occluded FAs did not display increased pain scores or deleterious effects on their gait or wellbeing.

Conclusion: Our data shows that sheep tolerate FA occlusion with no symptoms, similar to their cerebral circulation, making them an appropriate model for assessing small diameter femoral graft interposition studies and testing other cardiovascular interventions.

Autophagy is crucial for maintaining cellular homeostasis and is linked to various diseases. In Saccharomyces cerevisiae, the Polymyxin B Sensitivity 2 (Pbs2) protein is a member of the mitogen-activated protein kinase (MAPK) family and plays a role in mitophagy. To explore the potential role of Pbs2 in macroautophagy, we engineered wild-type and PBS2-deficient cells using plasmid construction and yeast transformation techniques, followed by a series of autophagy assays. First, after nitrogen starvation, the levels of autophagic activity were evaluated with the classical GFP-Atg8 cleavage assay and the Pho8Δ60 activity assay at different time points. Deleting PBS2 significantly decreased both GFP-Atg8 protein cleavage and Pho8Δ60 activity, indicating that Pbs2 is essential for macroautophagy. Furthermore, the influence of Pbs2 on macroautophagy was shown to be independent of Hog1, a well-known downstream factor of Pbs2. Second, the Atg8 lipidation assay demonstrated that Atg8 lipidation levels increased upon PBS2 deletion, suggesting that Pbs2 acts after Atg8 lipidation. Third, the proteinase K protection assay indicated that the loss of PBS2 led to a higher proportion of closed autophagosomes, implying that Pbs2 impacts the later stages of macroautophagy following autophagosome closure. In conclusion, Pbs2 regulates the late stages of macroautophagy induced by nitrogen starvation.

Tracheal collapse (TC), defined by excessive tracheal collapsibility, often results in severe respiratory distress in small-breed dogs. Surgical intervention, including the placement of extraluminal stents, has been employed as a treatment option. Owing to the anatomical and physiological similarities between rabbit and canine tracheas, a rabbit model was utilized to develop a novel extraluminal silicone tracheal stent and evaluate its feasibility in treating tracheomalacia. The stent was surgically implanted in eight New Zealand White rabbits after the induction of tracheomalacia. Postoperative evaluations, including clinical assessment, radiography, computed tomography (CT), and histological analysis, were performed at 1, 2, and 6 months post-implantation. All rabbits in the stent group survived without exhibiting signs of respiratory distress, whereas all rabbits in the tracheomalacia group experienced respiratory distress, with one succumbing to respiratory failure. Radiographic and CT evaluations confirmed that the stent effectively maintained airway patency, with tracheal measurements not significantly different from the preoperative values, indicating successful restoration of tracheal diameter. Histological analysis demonstrated minimal inflammatory response, the absence of fibrosis, and preserved structural integrity of the tracheal cartilage. Therefore, the novel extraluminal silicone tracheal stent provides effective airway support while minimizing adverse tissue reactions. Further studies, including the use of this stent in a canine TC model and assessment of its long-term outcomes, are warranted to explore its potential clinical applications in veterinary medicine.

The clinical application of hepatocyte transplantation has been significantly hindered by the scarcity of primary hepatocytes and the functional immaturity of in vitro–produced hepatocytes. By performing serial allogeneic hepatocyte transplantation in CRISPR/Cas9-mediated Fah-knockout pigs, we successfully achieved large-scale expansion of hepatocytes while maintaining their authentic biological characteristics. Particularly, the established model enables sustained in vivo liver reconstruction, concurrently ameliorating hepatic fibrosis and demonstrating functional microenvironmental remodeling. Moreover, through comprehensive single-cell transcriptomic profiling of 52 418 hepatocytes across transplant generations (F0–F2), we discovered that the cellular composition of these transplanted hepatocytes is similar to that of wild-type hepatocytes. The regenerated liver exhibits all six major hepatic cell types identical to the wild-type counterparts, with the characteristic lobular zonation patterns well preserved. Our research provides valuable insights into the large-scale expansion of physiologically functional hepatocytes in vivo without compromising their biological properties. This finding holds great promise for advancing the clinical application of human hepatocyte transplantation, potentially offering more effective treatment options for patients with liver diseases.