Search Results (119)

Metabolic dysfunction-associated steatotic liver disease (MASLD) is a widespread condition with a complex pathogenesis. Cell-based models are important tools for studying the mechanisms underlying its development and progression. The aim of this review is to analyze the HepaRG, Huh-7, immortalized human hepatocyte (IHH), and primary human hepatocyte (PHH) cell lines for modeling and studying MASLD. HepaRG represents the most metabolically competent immortalized hepatocyte model with preserved biotransformation activity and a physiological bioenergetic response to lipid loading, making it valuable for pharmacological and toxicological studies. Huh-7 is distinguished by its accessibility and suitability for studying steatosis, lipotoxicity, insulin resistance, and paracrine mechanisms of fibrogenesis; however, its use is limited by its tumor origin, impaired carbohydrate metabolism, and low activity of xenobiotic-metabolizing enzymes. The IHH model occupies an intermediate position because of its non-tumor origin and is of interest for studies of senescence, epigenetic regulation, and signaling pathways involved in steatosis, although interpretation of results requires consideration of immortalization-related effects and specific metabolic limitations. PHH remains the most physiologically relevant platform for MASLD modeling, particularly in three-dimensional (3D) and microphysiological formats; however, its use is limited by high cost, interindividual variability, and the limited duration of the differentiated phenotype. Increasing model complexity—from two-dimensional (2D) monocultures to co-cultures, spheroids, and organ-on-chip systems—enhances physiological relevance and enables reproduction not only of steatosis but also of the inflammatory and fibrogenic components of MASLD progression, yet it reduces reproducibility and complicates standardization. Overall, none of the existing models is universal, and the optimal strategy is to select models according to the specific research question. A key direction for future research is the standardization of steatosis induction protocols and the unification of criteria for evaluating results. Full article

Tripterygium glycosides extract (TGE), the primary active component of tripterygium glycosides tablets, is widely used for immune-related disorders but raises significant clinical concerns regarding cholestatic drug-induced liver injury. As conventional models fail to fully recapitulate the complex pathogenesis of traditional Chinese medicine toxicity, this study aimed to elucidate the mechanisms of TGE-induced cholestatic injury using a biomimetic microfluidic liver-on-a-chip platform. The chip integrated rat precision-cut liver slices (PCLSs) and human endothelial cells (EA.hy926) to simulate the hepatic sinusoidal microenvironment. Following TGE exposure (15–135 μg/mL for 12 and 24 h), vascular barrier integrity was maintained, while liver injury markers (ALT, AST, TBA, DBIL) significantly increased in a dose- and time-dependent manner, accompanied by progressive histopathological deterioration in PCLSs. Mechanistically, TGE triggered severe oxidative stress (decreased SOD/GSH/GSH-Px and increased MDA) and upregulated pro-inflammatory cytokines (IL-4 and IL-1β). Consequently, the expression of the bile acid receptor FXR and transporters (BSEP and MRP2) was significantly downregulated. In conclusion, TGE induces cholestatic liver injury via a sequential pathway: oxidative stress initiates an immune-inflammatory response, which subsequently suppresses the FXR/BSEP/MRP2 axis. Future studies should focus on developing fully humanized liver-on-a-chip systems to further validate these mechanisms and improve clinical translational significance. Full article

One of the common issues in the R&D of new drugs is the failure of clinical trials caused by the species-specific inadequacy of animal models to assess drugs’ efficiency and safety. Therefore, systems like organ-on-a-chip and, particularly, liver-on-a-chip (LOC) can be an efficient tool for recapitulating in vivo-like human physiology at the microscale. This review focuses on discussing LOC design, emphasizing its architecture and validation to reveal the trends in searching for a balance between biomimetics and functionality. We found that the huge variety of already published models can be divided into five groups based on their configuration complexity: flat one-channel, flat two-channel, vertically stacked multilayered, hexagonal-patterned, and multi-well chips. While researchers attempt to recapitulate the liver’s histology and its functions in detail by increasing the complexity of devices’ architectonics, industrial companies prefer to promote more simple and flexible solutions. Thus, the LOC designs of the future require neglecting some liver characteristics to make them standardizable and sustainable, which could facilitate their introduction into the market and clinics. Full article

Background and Objectives: Pro-inflammatory diet and high-fat diet (HFD) often coexist in real-world, but their combined impact on the gut–liver axis and potential nutritional countermeasures remain insufficiently studied. This study aimed to evaluate a pro-inflammatory–high-fat composite dietary pattern on the intestine and liver in the population, and to further evaluate the protective potential of astaxanthin (ATX) in complementary experimental systems. Methods: Data from the NHANES 2005–2010 were used to construct four composite exposure groups based on the dietary inflammation index (DII) and energy from fat. Survey-weighted regression analyses were performed to examine associations with systemic inflammation and liver injury. Interaction and C-reactive protein (CRP)-mediated effect analyses were conducted. Fifty SD rats were randomly divided into control group, model group induced by HFD combined with inflammatory factors, and low-, medium-, and high-dose Haematococcus pluvialis (HP) intervention groups. Serum lipids, liver enzymes, liver and colon pathology, and inflammatory and oxidative markers were measured in rats. In an in vitro organ-on-chip barrier model, the effect of ATX was observed when colonic barrier damage was induced using palmitic acid and lipopolysaccharides. Results: The high DII combined with HFD showed the largest increases in CRP, liver enzymes, and fatty liver index. A synergistic interaction was observed between DII and HFD, with CRP mediating approximately 20% of the effect. In rat model, HP-derived ATX improved the lipid profile, attenuated hepatic steatosis and oxidative damage, and reduced colonic pro-inflammatory cytokines, while restoration of tight junction proteins was limited. In colon organoid model, ATX showed limited efficacy in improving inflammation and barrier function. Conclusions: The pro-inflammatory–high-fat dietary pattern synergistically exacerbates gut–liver dysfunction. HP-derived ATX alleviates metabolic and inflammation-induced enterohepatic comorbidity, but its effect on repairing barrier structure is limited. Full article

Background: Huafeng Dan (HFD) is a traditional famous medicine from Guizhou Province, commonly used for the treatment of stroke-induced hemiplegia and epilepsy. Yaomu is a key component and serves as the sovereign herb in the formula. Most of the components of Yaomu are toxic Chinese herbal medicines. Traditional fermentation processing methods are required to reduce its toxicity. Purpose: Current studies have not yet systematically analyzed the chemical constituents before and after fermentation. Meanwhile, there is a lack of safety evaluation before and after the fermentation of Yaomu, which can provide a basis for safe clinical medication. Method: Chemical constituents of Yaomu before and after processing were analyzed using UHPLC-Q/TOF-MS to compare compositional changes induced by fermentation. To further screen potential toxic components, representative compounds were selected from these differential compounds based on statistical indicators (such as VIP value), low cost and easy availability, as well as criteria from the literature, and the content changes before and after fermentation were investigated. In vitro toxicity was evaluated using a microfluidic liver organ-on-a-chip model to assess the toxic effects of Yaomu extracts before and after fermentation. Results: Studies have shown that in both positive and negative ionization modes, a total of 361 compounds were annotated in unfermented Yaomu. After fermentation, a total of 350 compounds were annotated. Multivariate statistical analysis revealed significant differences in the chemical composition of Yaomu before and after fermentation. Quantitative analysis demonstrated that the levels of diester-type diterpenoid alkaloids were significantly reduced after fermentation, accompanied by concurrent decreases in lysophosphatidylcholine (LPC) species, compared with unfermented Yaomu. In contrast, the concentrations of amino alcohol-type diterpenoid alkaloids were significantly increased. The microfluidic liver organ-on-a-chip results demonstrated that the post-fermentation extract caused significantly attenuated impairment of hepatocellular function and viability. The in vitro toxicity findings showed good concordance. Full article

Microplastics (MPs) are increasingly recognized as a global environmental threat. Emerging evidence suggests they may have metabolic consequences. In this review, we synthesize current findings from animal and in vitro studies to propose a mechanistic framework linking MP exposure to type 2 diabetes mellitus (T2DM). This framework is uniquely centered on the gut–liver axis and macrophage-centric immune networks. We systematically delineate evidence suggesting that MPs can compromise intestinal barrier integrity, instigate gut dysbiosis, and promote pro-inflammatory M1 polarization of macrophages in experimental models. This immune activation is proposed to subsequently amplify hepatic inflammation, potentially contributing to systemic insulin resistance (IR) and pancreatic β-cell dysfunction. We emphasize that while this pathway is biologically plausible, direct causal evidence in humans remains limited and is a critical knowledge gap. Integrating multi-level evidence from animal models and in vitro systems, we delve into the trans-organ immunometabolic effects of MPs within adipose tissue, pancreas, and skeletal muscle, establishing their role as a novel class of “metabolic disruptors.” Critically, we assess the key controversies and knowledge gaps pertaining to dose–response relationships, particle-specific toxicity (size, polymer type, and additives), the effects of complex environmental mixtures, and the urgent need for robust human validation. We advocate for future research priorities, including multi-omics integration, advanced organ-on-a-chip platforms, prospective cohort studies, and targeted intervention strategies, to propel this field from mechanistic exploration toward clinical and public health relevance. Finally, this synthesis underscores that mitigating the production and environmental release of MPs, alongside developing strategies to impede their bioavailability and accumulation, represents a crucial public health imperative for the prevention of environment-related metabolic diseases. Full article

Enniatins (ENNs) are emerging Fusarium mycotoxins detected in food and feed. Despite their widespread occurrence, their toxicity remains poorly understood; thus, advanced in vitro systems that can mimic human physiology are of interest. We evaluated the cytotoxic and genotoxic effects of ENN A1 and ENN B1 exposure on differentiated (DIFF) and undifferentiated (UND) HepaRG liver cells cultured as 2D monolayers and 3D spheroids. Cytotoxicity, assessed by ATP-based luminescence, revealed a time-dependent decrease in inhibitory concentration 50 (IC₅₀) values between 24 h and 48 h across all models. In DIFF HepaRG cells, ENN A1 IC₅₀ values in 3D spheroids decreased from 14.4–18.2 µM at 24 h to 2.2–3.0 µM at 48 h, reaching values comparable to those measured in 2D DIFF cells at 48 h (2.2–2.6 µM), while no IC₅₀ could be determined in 2D at 24 h. For ENN B1, a pronounced time-dependent toxicity was observed, with IC₅₀ values in 3D DIFF spheroids decreasing from 4.1–6.6 µM at 24 h to 1.3–1.6 µM at 48 h, remaining lower than those measured in 2D DIFF cells at 48 h (2.4–3.0 µM). ENN A1 primarily induced apoptotic responses, whereas both ENN A1 and B1 were associated with necrotic responses, and ENN B1 induced a transient and limited autophagic signal, suggesting a minor role for autophagy. To further characterize cellular responses to ENN exposure, spheroids cultured in microfluidic chips were sectioned, and proliferation (Ki67), DNA damage (γH2AX), and apoptosis (cleaved caspase-3) was assessed. Immunostaining revealed no proliferative response, whereas significant DNA damage was detected, particularly in DIFF spheroids. At low, sub-cytotoxic concentrations (~5 µM, 24 h), ENN A1 induced significant DNA damage, as shown by increased γH2AX levels, while cytotoxic effects were only observed at higher concentrations (IC₅₀ ~ 18 µM, 24 h), supporting a potential genotoxic effect independent of cytotoxicity. Despite the structural similarities between ENN A1 and ENN B1, our results highlighted distinct cell death pathways between the two analogues. Both ENNs were detected throughout spheroids without evidence of peripheral restriction, although a homogeneous functional test could not be conclusively demonstrated. Overall, the 3D HepaRG spheroid model proved to be a more physiologically relevant system, offering differential sensitivity, as well as enhanced mechanistic insight, compared to 2D cultures. Full article

In vitro models for animal experiments serve as a crucial bridge connecting basic research and clinical translation, and their developmental history profoundly reflects the paradigm shifts in life science research. This article’s narrative reviews the evolutionary path from traditional two-dimensional (2D) cell culture to advanced three-dimensional (3D) organoid technology, focusing on how organoid technology overcomes the limitations of traditional models in terms of physiological relevance, species specificity, and ethical constraints. The review article elaborates on the current state of organoid research in veterinary science, including the construction of models for organs such as the intestine, liver, and reproductive system in livestock and companion animals. Addressing existing technical bottlenecks—such as insufficient model complexity, lack of standardization, and difficulties in simulating vascularization and the immune microenvironment—future development directions are proposed, including multi-organ chips, AI-assisted analysis, and the integration of gene editing. Research indicates that with the deep integration of cutting-edge technologies such as biomaterials, microfluidics, 3D printing, and AI, organoid technology is progressively becoming a core driver for advancing veterinary precision medicine, holding broad application prospects. Full article

Liver fibrosis is a significant consequence of severe liver injury resulting from viral hepatitis, alcohol, and metabolic dysfunction. Progressive fibrosis and ultimate cirrhosis are leading causes of morbidity and mortality worldwide, generally irreversible and poorly targeted by current therapies. Traditional in vitro models and animal models mostly fail to fully recapitulate human multicellular crosstalk, extracellular matrix (ECM) remodeling, and the chronic, immune modulated nature of the disease. Recent advances in three-dimensional (3D) cell culture models including organoids, spheroids, bioprinted constructs, and organ-on-a-chip systems are advantageous for reconstructing cellular diversity and mechanical microenvironments to understand pathophysiology and aid in drug discovery. Emerging multi-organ models are capable of incorporating microbiome derived cues and using multi-omics readouts and imaging-enabled mechanistic dissection for more predictive anti-fibrotic screening. These technologies align well with the recent Modernization 3.0 regulation and New Approach Methodologies by the Food and Drug Administration (FDA) and recent EU Pharmaceutical Reform. This review summarizes the pathophysiology of liver fibrosis, the current landscape of 3D human liver models, and examines how microbiome interfaces modulate fibrogenesis. Full article

The intestine–liver–muscle axis plays an essential role in drug and nutrient absorption, metabolism, and energy balance. Yet in vitro models capable of recapitulating this inter-organ communication remain limited. Here, we present a pump-free, 3D-printed multi-organ-on-a-chip device that enables dynamic co-culture of Caco-2 intestinal epithelial cells, HepG2 hepatocytes, and primary human skeletal myoblasts (HSkMs) under gravity-driven oscillatory flow. The device consists of five interconnected chambers designed to accommodate Transwell cell culture inserts for intestine and muscle compartments and hydrogel-embedded hepatocyte spheroids in the central hepatic compartment. The device was fabricated by low-cost fused deposition modeling (FDM) using acrylonitrile butadiene styrene (ABS) polymers. Under dynamic rocking, oscillatory perfusion promoted inter-organ communication without the need for external pumps or complex tubing. Biological assessments revealed that dynamic co-culture significantly enhanced the characteristics of skeletal muscle, as indicated by increased myosin heavy chain expression and elevated lactate production, while HepG2 spheroids exhibited improved hepatic function with higher albumin expression compared with monoculture. Additionally, Caco-2 cells maintained stable tight junctions and transepithelial electrical resistance, demonstrating preserved intestinal barrier integrity under dynamic flow. These results establish the device as a versatile, accessible 3D-printed platform for modeling the intestine–liver–muscle axis and investigating metabolic cross-talk in drug discovery and disease modeling. Full article

For more than a century, pathology has served as a cornerstone of modern medicine, relying primarily on static microscopic assessment of tissue morphology—such as H&E staining—which remains the “gold standard” for disease diagnosis. However, this conventional paradigm provides only a snapshot of disease states and often fails to capture their dynamic evolution and complex functional mechanisms. Moreover, animal models are constrained by marked interspecies differences, creating a persistent gap in translational research. To overcome these limitations, we propose the concept of New Pathophysiology, a research framework that transcends purely morphological descriptions and aims to resolve functional dynamics in real time. This approach integrates Organ-on-a-Chip (OOC) technology, multi-omics analyses, and artificial intelligence to reconstruct the entire course of disease initiation and to enable personalized medicine. In this review, we first outline the foundations and limitations of traditional pathology and animal models. We then systematically summarize more than one hundred existing OOC disease models across multiple organs—including the kidney, liver, and brain. Finally, we elaborate on how OOC technologies are reshaping the study of key pathological processes such as inflammation, metabolic dysregulation, and fibrosis by converting them into dynamic, mechanistic disease models, and we propose future perspectives in the field. This review adopts a relatively uncommon classification strategy based on pathological mechanisms (mechanism-based), rather than organ-based categorization, allowing readers to recognize shared principles underlying different diseases. Moreover, the focus of this work is not on emphasizing iteration or replacement of existing approaches, but on preserving past achievements from a historical perspective, with an emphasis on overcoming current limitations and enabling new advances. Full article

Microfluidic technologies have ushered in a new era in cardiac tissue engineering, providing more predictive in vitro models compared to two-dimensional culture studies. This review examines Organ-on-a-Chip (OoC) and Lab-on-a-Chip (LoC) platforms, with a specific focus on cardiovascular applications. OoCs, and particularly Heart-on-a-Chip systems, have advanced biomimicry to a higher level by recreating complex 3D cardiac microenvironments in vitro and dynamic fluid flow. These platforms employ induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs), engineered extracellular matrices, and dynamic mechanical and electrical stimulation to reproduce the structural and functional features of myocardial tissue. LoCs have introduced miniaturization and integration of analytical functions into compact devices, enabling high-throughput screening, advanced diagnostics, and efficient pharmacological testing. They enable the investigation of pathophysiological mechanisms, the assessment of cardiotoxicity, and the development of precision medicine approaches. Furthermore, progress in multi-organ systems expands the potential of microfluidic technologies to simulate heart–liver, heart–kidney, and heart–tumor interactions, providing more comprehensive predictive models. However, challenges remain, including the immaturity of iPSC-derived cells, the lack of standardization, and scalability issues. In general, microfluidic platforms represent strategic tools for advancing cardiovascular research in translation and accelerating therapeutic innovation within precision medicine. Full article

Malaria remains one of the devastating illnesses, and drug-resistant malaria has incurred enormous societal costs. A few host kinases are vital for the liver stage malaria and might be promising drug targets against drug-resistant malaria. STK35L1 is one of the host kinases that is highly upregulated during the liver stage of malaria, and the knockdown of STK35L1 significantly suppresses Plasmodium sporozoite infection. In this study, we retrieved the promoter region of STK35L1 based on 5′ complete transcripts, transcription start sites, and cap analysis of gene expression tags. Furthermore, we identify transcriptionally active regions by analyzing CpG islands, histone acetylation (H3K27ac), and histone methylation (H3K4me3). It suggests that the identified promoter region is active and has cis-regulatory elements and enhancer regions. We identified various putative transcription factors (TFs) from the various high-throughput ChIP data that might bind to the promoter region of STK35L1. These TFs were differentially regulated during the infection of Plasmodium sporozoites in HepG2 cells. Our molecular modeling study suggests that, except for SMAD3, the identified TFs may be directly bound to the promoter. Together, the data suggest that these TFs may play a role in sporozoite infection and in regulating STK35L1 expression during the liver stage of malaria. Full article

Endothelial-to-mesenchymal transition (EndMT) is a cellular program implicated in fibrosis, vascular remodeling, and the tumor microenvironment across multiple organs. We synthesize mechanistic pathways including TGF-β/SMAD, non-canonical (MAPK, PI3K/AKT, Rho/ROCK), Notch, and Wnt/β-catenin cascades. Their crosstalk with hypoxia, inflammatory cues, and epigenetic mechanisms can drive loss of endothelial identity and acquisition of mesenchymal characteristics. We outline disease contexts in the heart, lungs, kidneys, liver, central nervous system, and cancer, highlighting context-dependent contributory roles of EndMT. Therapeutically, we review pathway-targeted agents, epigenetic inhibitors, microRNA-based strategies, antibodies/biologics, small molecules and natural compounds, and cell- and gene-based interventions. Finally, we outline a translational roadmap that pairs patient-derived iPSC/organoid and organ-on-a-chip platforms to stratify EndMT states and prioritize targets. We also explore combination regimens that integrate multi-pathway modulation with epigenetic and immune approaches, aiming to deliver clinically meaningful anti-fibrotic benefits while better preserving physiological signaling. Full article

Organic anion transporting polypeptide 1B1 (OATP1B1) is specifically expressed at the basolateral membrane of human liver cells and transports a wide range of endogenous compounds, toxins, and drugs, making it a crucial factor in determining the pharmacokinetics of many clinically important medications. Triptergium wilfordii Hook. f. (TWHF) is a traditional Chinese medicine known for its long history of therapeutic effects. A previous study conducted in our laboratory found that major components of TWHF, including wilforine (WFR), wilforgine (WFG), celastrol (CL), and triptolide (TPL), directly suppressed the function of OATP1B1. In the current study, we investigated the long-term (24 h) effects of these TWHF components on the transporter. It was found that TPL was the most potent compound exhibiting inhibitory effects. Mechanistically, TPL accelerated the degradation of OATP1B1, which is likely mediated by serum and glucocorticoid-induced kinase 1 (SGK1). TPL downregulated the mRNA expression of SGK1 and reduced the nuclear accumulation of nuclear factor kappa B (NFκB). Further analysis of the upstream sequence of SGK1 identified three potential binding sites for NFκB. Both luciferase activity assays and chromatin immunoprecipitation (ChIP) analyses confirmed the binding of NFκB to two specific sites located at −1015 bp~−1006 bp and −319 bp~−310 bp. Full article