Rheumatoid arthritis (RA) is a chronic autoimmune disease that affects approximately 0.46% of the global population. Conventional therapeutics for RA, including disease-modifying antirheumatic drugs (DMARDs), nonsteroidal anti-inflammatory drugs (NSAIDs), and corticosteroids, frequently result in unintended adverse effects. Dexamethasone (DEX) is a potent glucocorticoid used to treat RA due to its anti-inflammatory and immunosuppressive properties. Liposomal delivery of DEX, particularly when liposomes are surface-modified with targeting ligands like peptides or sialic acid, can improve drug efficacy by enhancing its distribution to inflamed joints and minimizing toxicity. This study investigates the potential of liposomal drug delivery systems to enhance the efficacy and targeting of DEX in the treatment of RA. Results from various studies demonstrate that liposomal DEX significantly inhibits arthritis progression in animal models, reduces joint inflammation and damage, and alleviates cartilage destruction compared to free DEX. The liposomal formulation also shows better hemocompatibility, fewer adverse effects on body weight and immune organ index, and a longer circulation time with higher bioavailability. The anti-inflammatory mechanism is associated with the downregulation of pro-inflammatory cytokines like tumor necrosis factor-α (TNF-α) and B-cell–activating factor (BAFF), which are key players in the pathogenesis of RA. Additionally, liposomal DEX can induce the expression of anti-inflammatory cytokines like interleukin-10 (IL-10), which has significant anti-inflammatory and immunoregulatory properties. The findings suggest that liposomal DEX represents a promising candidate for effective and safe RA therapy, with the potential to improve the management of this debilitating disease by providing targeted delivery and sustained release of the drug.
Human-derived tumor models are essential for preclinical development of new anticancer drug entities. Generating animal models bearing tumors of human origin, such as patient-derived or cell line–derived xenograft tumors, is dependent on immunodeficient strains. Tumor-bearing immunodeficient mice are susceptible to developing unwanted disorders primarily irrelevant to the tumor nature; and if get involved with such disorders, reliability of the study results will be undermined, inevitably confounding the research in general. Therefore, a rigorous health surveillance and clinical monitoring system, along with the establishment of a strictly controlled barrier facility to maintain a pathogen-free state, are mandatory. Even if all pathogen control and biosafety measures are followed, there are various noninfectious disorders capable of causing tissue and multiorgan damage in immunodeficient animals. Therefore, the researchers should be aware of sentinel signs to carefully monitor and impartially report them. This re discusses clinical signs of common unwanted disorders in experimental immunodeficient mice, and how to examine and report them.
Background: Dengue fever, an acute insect-borne infectious disease caused by the dengue virus (DENV), poses a great challenge to global public health. Hepatic involvement is the most common complication of severe dengue and is closely related to the occurrence and development of disease. However, the features of adaptive immune responses associated with liver injury in severe dengue are not clear.
Methods: We used single-cell sequencing to examine the liver tissues of mild or severe dengue mice model to analyze the changes in immune response of T cells in the liver after dengue virus infection, and the immune interaction between macrophages and T cells. Flow cytometry was used to detect T cells and macrophages in mouse liver and blood to verify the single-cell sequencing results.
Results: Our result showed CTLs were significantly activated in the severe liver injury group but the immune function-related signal pathway was down-regulated. The reason may be that the excessive immune response in the severe group at the late stage of DENV infection induces the polarization of macrophages into M2 type, and the macrophages then inhibit T cell immunity through the TGF-β signaling pathway. In addition, the increased proportion of Treg cells suggested that Th17/Treg homeostasis was disrupted in the livers of severe liver injury mice.
Conclusions: In this study, single-cell sequencing and flow cytometry revealed the characteristic changes of T cell immune response and the role of macrophages in the liver of severe dengue fever mice. Our study provides a better understanding of the pathogenesis of liver injury in dengue fever patients.
Background: To study the relationships among emodin, synovial fibroblasts (FLSs), and macrophages (STMs) to provide guidance for the use of emodin in rheumatoid arthritis (RA) treatment.
Methods: RA clinical samples from patients with different pathological processes were collected, and the correlations between the subsets of FLSs and STMs and pathological processes were analyzed via flow cytometry. In vitro experimental methods such as enzyme linked immunosorbent assay (ELISA), Western blotting, Transwell assays, CCK-8 assays and cell coculture were used to assess cell proliferation, migration and secretion of inflammatory factors. A collagen-induced arthritis mouse model was constructed to investigate the therapeutic potential of emodin in RA by flow cytometry, micro-CT and staining.
Results: Unique subsets of FLSs and STMs, namely, FAPα+THY1- FLSs, FAPα+THY1+ FLSs, and MerTKposTREM2high STMs, were identified in synovial tissues from RA patients. The number of MerTKposTREM2high STMs was negatively correlated with the degree of damage in RA, while the number of FAPα+THY1- FLSs was positively correlated with damage. On the one hand, emodin promoted the aggregation of MerTKposTREM2high STMs. Moreover, MerTKposTREM2high STM-mediated secretion of exosomes was promoted, which can inhibit the secretion of pro-inflammatory factors by FAPα+THY1+ FLSs and promote the secretion of anti-inflammatory factors by FAPα+THY1+ FLSs, thereby inhibiting FAPα+THY1-FLS proliferation and migration, improving the local immune microenvironment, and inhibiting RA damage.
Conclusion: Emodin was shown to regulate the aggregation of STM subsets and exosome secretion, affecting the secretion, proliferation and migration of inflammatory factors in FLS subsets, and ultimately achieving good therapeutic efficacy in RA patients, suggesting that it has important clinical value.
Background: Macrophages are the primary innate immune cells encountered by the invading coronaviruses, and their abilities to initiate inflammatory reactions, to maintain the immunity homeostasis by differential polarization, to train the innate immune system by epigenic modification have been reported in laboratory animal research.
Methods: In the current in vitro research, murine macrophage RAW 264.7 cell were infected by mouse hepatitis virus, a coronavirus existed in mouse. At 3-, 6-, 12-, 24-, and 48-h post infection (hpi.), the attached cells were washed with PBS and harvested in Trizol reagent. Then The harvest is subjected to transcriptome sequencing.
Results: The transcriptome analysis showed the immediate (3 hpi.) up regulation of DEGs related to inflammation, like Il1b and Il6. DEGs related to M2 differential polarization, like Irf4 showed up regulation at 24 hpi., the late term after viral infection. In addition, DEGs related to metabolism and histone modification, like Ezh2 were detected, which might correlate with the trained immunity of macrophages.
Conclusions: The current in vitro viral infection study showed the key innated immunity character of macrophages, which suggested the replacement value of viral infection cells model, to reduce the animal usage in preclinical research.
Cardiac injury initiates repair mechanisms and results in cardiac remodeling and fibrosis, which appears to be a leading cause of cardiovascular diseases. Cardiac fibrosis is characterized by the accumulation of extracellular matrix proteins, mainly collagen in the cardiac interstitium. Many experimental studies have demonstrated that fibrotic injury in the heart is reversible; therefore, it is vital to understand different molecular mechanisms that are involved in the initiation, progression, and resolution of cardiac fibrosis to enable the development of antifibrotic agents. Of the many experimental models, one of the recent models that has gained renewed interest is isoproterenol (ISP)–induced cardiac fibrosis. ISP is a synthetic catecholamine, sympathomimetic, and nonselective β-adrenergic receptor agonist. The overstimulated and sustained activation of β-adrenergic receptors has been reported to induce biochemical and physiological alterations and ultimately result in cardiac remodeling. ISP has been used for decades to induce acute myocardial infarction. However, the use of low doses and chronic administration of ISP have been shown to induce cardiac fibrosis; this practice has increased in recent years. Intraperitoneal or subcutaneous ISP has been widely used in preclinical studies to induce cardiac remodeling manifested by fibrosis and hypertrophy. The induced oxidative stress with subsequent perturbations in cellular signaling cades through triggering the release of free radicals is considered the initiating mechanism of myocardial fibrosis. ISP is consistently used to induce fibrosis in laboratory animals and in cardiomyocytes isolated from animals. In recent years, numerous phytochemicals and synthetic molecules have been evaluated in ISP-induced cardiac fibrosis. The present review exclusively provides a comprehensive summary of the pathological biochemical, histological, and molecular mechanisms of ISP in inducing cardiac fibrosis and hypertrophy. It also summarizes the application of this experimental model in the therapeutic evaluation of natural as well as synthetic compounds to demonstrate their potential in mitigating myocardial fibrosis and hypertrophy.
Frozen shoulder (FS), also known as adhesive capsulitis, is a condition that causes contraction and stiffness of the shoulder joint capsule. The main symptoms are persistent shoulder pain and a limited range of motion in all directions. These symptoms and poor prognosis affect people’s physical health and quality of life. Currently, the specific mechanisms of FS remain unclear, and there is variability in treatment methods and their efficacy. Additionally, the early symptoms of FS are difficult to distinguish from those of other shoulder diseases, complicating early diagnosis and treatment. Therefore, it is necessary to develop and utilize animal models to understand the pathogenesis of FS and to explore treatment strategies, providing insights into the prevention and treatment of human FS. This paper res the rat models available for FS research, including external immobilization models, surgical internal immobilization models, injection modeling models, and endocrine modeling models. It introduces the basic procedures for these models and compares and analyzes the advantages, disadvantages, and applicability of each modeling method. Finally, our paper summarizes the common methods for evaluating FS rat models.
Background: Apoptosis signal-regulating kinase 1 (ASK1) is a MAP3K kinase in the MAPK signaling pathway activated by stressors and triggers downstream biological effects such as inflammation and apoptosis; therefore, inhibition of ASK1 kinase activity can protect cells from pathological injury. In this study, we designed and synthesized a novel selective ASK1 inhibitor, CS17919, and investigated its pharmacological effects in various animal models of metabolic injury.
Methods: First, we validated the ability of CS17919 to inhibit ASK1 in vitro and then tested the safety profile of CS17919 in cell lines compared with Selonsertib (GS-4997), a phase III ASK1 inhibitor. We then conducted pharmacokinetic (PK) studies in mice. Finally, we tested the in vivo efficacy of CS17919 in murine models of chronic kidney disease (CKD) and non-alcoholic steatohepatitis (NASH).
Results: Compared to GS-4997, CS17919 demonstrated comparable inhibition of ASK1 in vitro, exhibited lower toxicity, and provided greater protection in palmitic acid-treated LO2 cells. CS17919 also showed pronounced pharmacokinetic properties such as a high plasma concentration. In the unilateral ureteral obstruction model (UUO), CS17919 and GS-4997 preserved kidney function and showed a non-significant tendency to alleviate kidney fibrosis. In the diabetic kidney disease (DKD) model, CS17919 significantly improved serum creatinine and glomerular sclerosis. In the NASH model, the combination of CS17919 and a THRβ agonist (CS27109) was found to significantly improve liver inflammation and substantially reduced liver fibrosis.
Conclusions: CS17919 showed cell protective, anti-inflammatory, and antifibrotic effects in vitro and in vivo, suggesting its therapeutic potential for metabolic-related kidney and liver diseases.
Background: The emerging incidence of pathogenic liver conditions is turning into a major concern for global health. Induction of pyroptosis in hepatocytes instigates cellular disintegration, which in turn liberates substantial quantities of pro-inflammatory intracellular substances, thereby accelerating the advancement of liver fibrosis. Consequently, directing therapeutic efforts towards inhibiting pyroptosis could potentially serve as an innovative approach in managing inflammation related chronic hepatic disorders.
Methods: GSDMD-NTki/wt mice and Alb-creki/wt mice were generated using CRISPR/9 technology. After crossing the two strains together, we induced conditional cell death by doxycycline to construct a mouse model of liver fibrosis. We analyzed differentially expressed genes by RNA sequencing and explored their biological functions. The efficacy of obeticholic acid (OCA) in the treatment of liver fibrosis was assessed.
Results: Doxycycline-treated GSDMD-NTki/wt×Alb-creki/wt mice showed severe liver damage, vacuolation of hepatocytes, increased collagen fibers, and accumulation of lipid droplets. The expression of liver fibrosis related genes was greatly increased in the doxycycline-treated mouse liver compared with untreated mouse liver. RNA-sequencing showed that upregulated differentially expressed genes were involved in inflammatory responses, cell activation, and metabolic processes. Treatment with OCA alleviated the liver fibrosis, with reduced ALT and AST levels seen in the GSDMD-NTki/wt×Alb-creki/wt mice.
Conclusions: We successfully constructed a novel mouse model for liver fibrosis. This GSDMD-NT-induced fibrosis may be mediated by abnormal lipid metabolism. Our results demonstrated that we successfully constructed a mouse model of liver fibrosis, and GSDMD-NT induced fibrosis by mediating lipid metabolism.
Background: Over the past few decades, a threefold increase in obesity and type 2 diabetes (T2D) has placed a heavy burden on the health-care system and society. Previous studies have shown correlations between obesity, T2D, and neurodegenerative diseases, including dementia. It is imperative to further understand the relationship between obesity, T2D, and cognitive deficits.
Methods: This investigation tested and evaluated the cognitive impact of obesity and T2D induced by high-fat diet (HFD) and the effect of the host genetic background on the severity of cognitive decline caused by obesity and T2D in collaborative cross (CC) mice. The CC mice are a genetically diverse panel derived from eight inbred strains.
Results: Our findings demonstrated significant variations in the recorded phenotypes across different CC lines compared to the reference mouse line, C57BL/6J. CC037 line exhibited a substantial increase in body weight on HFD, whereas line CC005 exhibited differing responses based on sex. Glucose tolerance tests revealed significant variations, with some lines like CC005 showing a marked increase in area under the curve (AUC) values on HFD. Organ weights, including brain, spleen, liver, and kidney, varied significantly among the lines and sexes in response to HFD. Behavioral tests using the Morris water maze indicated that cognitive performance was differentially affected by diet and genetic background.
Conclusions: Our study establishes a foundation for future quantitative trait loci mapping using CC lines and identifying genes underlying the comorbidity of Alzheimer’s disease (AD), caused by obesity and T2D. The genetic components may offer new tools for early prediction and prevention.
Background: We investigated the similarities and differences between two experimental approaches using tachy-pacing technology to induce desynchronized heart failure in canines.
Methods: A total of eight dogs were included in the experiment, four were tachy-paced in right ventricle apex (RVAP) and 4 were paced in right atrium after the ablation of left bundle branch to achieve left bundle branch block (RAP+LBBB). Three weeks of follow-up were conducted to observe the changes in cardiac function and myocardial staining was performed at the end of the experiment.
Results: Both experimental approaches successfully established heart failure with reduced ejection fraction models, with similar trends in declining cardiac function. The RAP+LBBB group exhibited a prolonged overall ventricular activation time, delayed left ventricular activation, and lesser impact on the right ventricle. The RVAP approach led to a reduction in overall right ventricular compliance and right ventricular enlargement. The RAP+LBBB group exhibited significant reductions in left heart compliance (LVGLS, %: RAP+LBBB -12.60±0.12 to -5.93±1.25; RVAP -13.28±0.62 to -8.05±0.63, p=0.023; LASct, %: RAP+LBBB -15.75±6.85 to -1.50±1.00; RVAP -15.75±2.87 to -10.05±6.16, p=0.035). Histological examination revealed more pronounced fibrosis in the left ventricular wall and left atrium in the RAP+LBBB group while the RVAP group showed more prominent fibrosis in the right ventricular myocardium.
Conclusion: Both approaches establish HFrEF models with comparable trends. The RVAP group shows impaired right ventricular function, while the RAP+LBBB group exhibits more severe decreased compliance and fibrosis in left ventricle.
Background: The aim of this study was to analyze the bi-directional causal relationship between lipid profile and characteristics related to muscle atrophy by using a bi-directional Mendelian randomization (MR) analysis.
Methods: The appendicular lean mass (ALM), whole body fat-free mass (WBFFM) and trunk fat-free mass (TFFM) were used as genome-wide association study (GWAS) data for evaluating muscle mass; the usual walking pace (UWP) and low grip strength (LGS) were used as GWAS data for evaluating muscle strength; and the triglycerides (TG), total cholesterol (TC), high density lipoprotein cholesterol (HDL), low density lipoprotein cholesterol (LDL), apolipoprotein A-1 (Apo A-1), and apolipoprotein B (Apo B) were used as GWAS data for evaluating lipid profile. For specific investigations, we mainly employed inverse variance weighting for causal estimation and MR-Egger for pleiotropy analysis.
Results: MR results showed that the lipid profile predicted by genetic variants was negatively correlated with muscle mass, positively correlated with UWP, and was not causally correlated with LGS. On the other hand, the muscle mass predicted by genetic variants was negatively correlated with lipid profile, the UWP predicted by genetic variants was mainly positively correlated with lipid profile, while the LGS predicted by genetic variants had no relevant causal relationship with lipid profile.
Conclusions: Findings of this MR analysis suggest that hyperlipidemia may affect muscle mass and lead to muscle atrophy, but has no significant effect on muscle strength. On the other hand, increased muscle mass may reduce the incidence of dyslipidemia.
Background: Breast cancer is the most common cancer in women, and in advanced stages, it often metastasizes to the brain. However, research on the biological mechanisms of breast cancer brain metastasis and potential therapeutic targets are limited.
Methods: Differential gene expression analysis (DEGs) for the datasets GSE43837 and GSE125989 from the GEO database was performed using online analysis tools such as GEO2R and Sangerbox. Further investigation related to SULF1 was conducted using online databases such as Kaplan–Meier Plotter and cBioPortal. Thus, expression levels, variations, associations with HER2, biological processes, and pathways involving SULF1 could be analyzed using UALCAN, cBioPortal, GEPIA2, and LinkedOmics databases. Moreover, the sensitivity of SULF1 to existing drugs was explored using drug databases such as RNAactDrug and CADSP.
Results: High expression of SULF1 was associated with poor prognosis in advanced breast cancer brain metastasis and was positively correlated with the expression of HER2. In the metastatic breast cancer population, SULF1 ranked top among the 16 DEGs with the highest mutation rate, reaching 11%, primarily due to amplification. KEGG and GSEA analyses revealed that the genes co-expressed with SULF1 were positively enriched in the ‘ECM-receptor interaction’ gene set and negatively enriched in the ‘Ribosome’ gene set. Currently, docetaxel and vinorelbine can act as treatment options if the expression of SULF1 is high.
Conclusions: This study, through bioinformatics analysis, unveiled SULF1 as a potential target for treating breast cancer brain metastasis (BM).
The present study aims to establish a reproducible large animal experimental unit using a minipig model to monitor cardiac function changes. A 90-min closed-chest balloon occlusion of the left anterior descending branch of the coronary artery was used to induce myocardial infarction in Pannon minipigs. To monitor the cardiac function, measurements were made by cardiac magnetic resonance imaging (cMRI), invasive pressure monitoring, and a Pulse index Continuous Cardiac Output (PiCCO) hemodynamic system at 0, 72, and 720 h during the follow-up period. End-diastolic and end-systolic volumes (EDV, ESV), left ventricular ejection fraction (LVEF) obtained by cMRI evaluation, global ejection fraction and aortic dP/dtmax obtained by the invasive method, were recorded and compared. The 72- and 720-h EDV data showed a significant increase (p=0.012, <0.001) compared to baseline, and the Day 30 data showed a significant increase compared to Day 3 (p=0.022). The ESV 72 h after the infarction showed a significant increase (p=0.001) compared to baseline, which did not change significantly by Day 30 (p=0.781) compared to Day 3. EDV and ESV were significantly negatively correlated with aortic dpmax, and ESV was significantly correlated with LVEF. For LVEF and dPmax, a significant (p<0.001 and p=0.002) worsening was demonstrated at Day 3 compared to baseline, which was no longer statistically detectable for LVEF at Day 30 (p=0.141), while the difference for dPmax was maintained (p=0.002). The complementary use of PiCCO hemodynamic measurements in large animal models makes the previously used methodologies more robust and reliable.
Background: Previously, a chronic social defeat stress (CSDS) model has been widely-adopted for assessing depressive-like behaviors in animals. However, there is still room for improvement in the CSDS model to safeguard study accuracy and the welfare of lab rodents. Our study team developed a novel, standardized apparatus to induce CSDS in rodents and assessed the model’s practical adaptability.
Methods: An innovative CSDS cage apparatus and water bottle was designed. To evaluate the effectiveness of the newly developed tools, a variety of animal models, including the tail suspension test (TST), sucrose preference test, forced swimming test (FST), novelty-suppressed feeding test, female urine sniffing test, and open field test (OFT), were adopted to assess depressive-like behaviors in mice. Fluoxetine treatment was also administered to observe the reversal effect, as part of the validation.
Results: The CSDS cage apparatus resulted in the manifestation of depressive-like behaviors in the model mice. Significant reductions in sucrose preference and urine sniffing time were observed, while the OFT revealed decreased central zone total distance, residence time, and frequency of entry. Moreover, increased immobility was found in the FST and TST. Fluoxetine treatment was found to successfully reverse the modeling effect.
Conclusion: The CSDS cage apparatus was validated for enhanced usability and addressed the previous challenges of water bottle leakage and lab rodent welfare issues. The consistent results from multiple behavioral tests also supported real-world application of the apparatus, offering researchers a promising alternative to conventional rodent cages.
Background: Traditional DNA microinjection methods used in mammals are difficult to apply to avian species due to their unique reproductive characteristics. Genetic manipulation in chickens, particularly involving immature follicles within living ovaries, has not been extensively explored. This study seeks to establish an efficient method for generating transgenic chickens through ovarian injection, potentially bypassing the challenges associated with primordial germ cell (PGC) manipulation and fertilized egg microinjection.
Methods: Hens were anesthetized and underwent a surgical procedure to access the ovary for DNA injection into immature follicles. The study used liposomes to deliver GFP-expressing plasmids at various dosages. After injection, hens recovered, and their eggs were fertilized through artificial insemination.
Results: Transgenic chickens were successfully generated in one generation without needing G0 founders. The injection of 20 µg plasmid yielded the highest transgenic efficiency at 12.1%. GFP-positive embryos were confirmed through microscopy, and successful transgene expression was validated at the tissue level using immunostaining. TERT and GFP elements introduced in the G1 generation resulted in 4.1% positive transgene rates, as confirmed by PCR and Southern blotting.
Conclusion: This ovarian injection method offers a promising alternative for avian genetic manipulation, bypassing complex PGC procedures and enabling direct generation of G1 transgenic chickens. This technique simplifies the transgenic process for chickens and has the potential to be adapted for other avian species, especially those without established PGCs culture systems.