Experimental Animals最新文献

Calcium/calmodulin-dependent protein kinase II (CAMKII) is a critical regulator of cardiac electrophysiology. However, the role of the four bases deletion polymorphism in Camk2d which codes delta subunit of CAMKII, particularly those involving intron sequences, remains poorly understood. This study aimed to investigate the impact of Camk2d c.1044+125_128delGTTT missing polymorphism on cardiac morphology and arrhythmogenesis in normal adult Sprague-Dawley (SD) rats. A total of 85 SD rats were genotyped by Sanger sequencing, revealing a distribution of 25.9% wild-type (WT), 48.2% heterozygous, and 25.9% homozygous variants. Echocardiography, Hematoxylin-Eosin staining, Masson's trichrome staining and transmission electron microscopy indicated no significant differences in cardiac structure or baseline function among the three groups. In freely moving rats, premature atrial arrhythmias were detected in 2 of 9 WT rats, 1 of 9 heterozygous rats, and 1 of 9 homozygous rats. Premature ventricular contractions (PVCs) were observed in none of 9 WT or homozygous rats, 3 of 9 heterozygous rats, with one heterozygous rat exhibiting frequent PVCs. Electrical programmed stimulation revealed a higher incidence of inducible atrial fibrillation in homozygous rats compared to WT rats and a higher incidence of inducible ventricular tachycardia in heterozygous rats compared to WT rats. These findings suggest that deletion polymorphism in the intron sequences of Camk2d are unexpectedly common in normal SD rat populations and that such polymorphism predispose to ventricular arrhythmias without overt structural heart disease. Our study highlights the potential arrhythmogenic risk associated with non-coding DNA sequence alterations in Camk2d and underscores the importance of genetic screening in experimental animal models.

Endothelial dysfunction (ED) plays a pivotal role in the pathogenesis of hypertension and its associated vascular complications. Qingda granule (QDG) exhibits significant antihypertensive properties and demonstrates therapeutic potential in ameliorating vascular dysfunction. This study aimed to explore QDG's role in alleviating endothelial injury in hypertension. An L-NAME (Nω-Nitro-L-arginine methyl ester)-induced hypertensive mouse model was used to evaluate the effects of QDG on blood pressure and endothelial function. Endothelial function was assessed through histological analysis, nitric oxide (NO) quantification, and vascular response measurements. To explore underlying mechanisms, network pharmacology was conducted using databases such as HERB, SwissTargetPrediction and STRING. Key pathways related to inflammation and cell adhesion were identified. Based on these findings, immunohistochemical staining was conducted to analyze the expression of phosphorylation of NF-kappaB p65 (p-NF-κB p65), NF-κB p65, intercellular adhesion molecule-1 (ICAM-1), and tumor necrosis factor-α (TNF-α) in vascular tissues. QDG treatment significantly reduced blood pressure, increased NO levels, and enhanced endothelial nitric oxide synthase (eNOS) expression in L-NAME-induced hypertensive mice, indicating its potential to restore endothelial function. Experimental validation further confirmed that QDG markedly suppressed the expression of p-NF-κB p65, TNF-α, and ICAM-1 in vascular tissues. These results suggest that QDG alleviates hypertension-induced ED primarily by inhibiting inflammation and endothelial adhesion via the NF-κB signaling pathway. Overall, QDG presents a promising therapeutic candidate for managing hypertension and its vascular complications.

In pharmaceutical development, weight loss is occasionally observed in monkeys during non-clinical toxicity studies and can be difficult to differentiate from drug effects. This study retrospectively analyzed data from control group monkeys without drug treatment to investigate the incidence of weight loss and its physiological and pathological characteristics. We also investigated potential improvements through enhanced animal welfare. In the 4- and 13-week toxicity studies conducted at the test facility from 2010 to 2022, 684 control group monkeys were investigated. Among them, 3 animals in the 4-week toxicity studies and 5 animals in the 13-week toxicity studies showed a weight change rate of less than -10%, resulting in an incidence rate of 1.2%. However, these animals had adequate food consumption. Animals in the 4-week toxicity studies showed signs of stress in histopathology. Additionally, 2/3 animals in the 4-week toxicity studies had decreased blood glucose levels and 1/5 animal in the 13-week toxicity study fell into a crouching posture, suggesting hypoglycemia that was alleviated with glucose administration, indicating stress-induced metabolic abnormalities. From 2015, an enrichment program was implemented to improve animal welfare. Prior to this program, 2.4% of animals showed a weight change rate of less than -10%, which dropped to 0.25% post-implementation, suggesting the program's effectiveness in reducing stress. These results clarify the characteristics of animals that lose weight during toxicity studies and suggest that improving animal welfare can reduce the incidence rate.

Allergic rhinitis (AR) is an inflammatory disorder driven primarily by aberrant T helper 2 (Th2) differentiation in CD4⁺ T cells. Although dual-specificity phosphatase 5 (DUSP5) has been implicated in inflammatory and autoimmune regulation, its role in AR remains unexplored. In this study, an AR mouse model was established via intraperitoneal sensitization and intranasal challenge with ovalbumin. We observed significant downregulation of DUSP5 expression in the nasal mucosa, particularly within CD4⁺ cells. To elucidate its function, a lentiviral vector overexpressing DUSP5 was constructed and used to transduce naive CD4⁺ T cells isolated from BALB/c mouse spleens. Overexpression of DUSP5 suppressed Th2-specific cytokine production and inhibited Th2 differentiation. Mechanistic investigations using a luciferase reporter assay revealed that Dusp5 is transcriptionally repressed by SRY-box transcription factor 11 (SOX11), a known transcription factor that promotes the progression of AR. Furthermore, DUSP5 overexpression counteracted the pro-Th2 effects mediated by SOX11. These results demonstrate that DUSP5, transcriptionally inhibited by SOX11, attenuates AR-associated inflammation by restraining Th2 differentiation. Our findings identify DUSP5 as a potential therapeutic target for AR.

DNA double-strand breaks (DSBs) are among the most hazardous cellular damages, potentially leading to cell death or oncogenesis if unrepaired. Genome editing methods, such as the CRISPR/Cas9 system, induce DSBs and utilize these repair pathways for gene knockout and knock-in. Although ionizing radiation also induces DSBs, it is not clear whether the efficiency of genome editing is affected by ionizing radiation. This study investigated the impact of gamma-ray exposure on the genome editing efficiency of the improved genome editing via oviductal nucleic acid delivery (i-GONAD) method. Gamma-rays were exposed to pregnant mice receiving i-GONAD targeting the Hr gene, whose mutation causes hair loss in mice. The exposure on the fertilization day (Day 0) decreased natural delivery rates and litter sizes, with notable effects at 0.3 Gy or higher. Although the proportions of hairless offspring obtained by i-GONAD differed greatly between single-guide RNAs (sgRNAs) used, total mutation rates, including hairless, mosaic, and indel, were equivalent. Gamma-ray exposure on Day 0 and the day after fertilization (Day 1) similarly and almost dose-dependently enhanced the genome editing efficiency evaluated by the total mutation rate. This study suggests the improvement of genome editing efficiency by gamma-ray exposure, at least in i-GONAD method, potentially facilitating the creation of diverse experimental animal models.

Chronic obstructive pulmonary disease (COPD) is a prevalent lung disease that mainly induced by cigarette smoking (CS). Soyasaponin I is an amphiphilic oleanane triterpenoid glycoside extracted form Astragali Radix. In order to investigate treatment strategies of COPD, this study focused on the effect of soyasaponin I on the lung tissue of COPD model. The mouse model of COPD was induced by CS exposure for 12 weeks, and was administrated with different doses of soyasaponin I. Subsequently, the morphology and histopathology of lung tissue, the proportion of inflammatory cell, the levels of inflammatory cytokines, and indicators of oxidative stress were assessed and analyzed. The signaling pathway potentially regulated by soyasaponin I in the pathogenesis of COPD were predicted by network pharmacology analysis and validated by western blot. Our results demonstrated that soyasaponin I mitigated the lung injury and bronchial lesions induced by COPD through reducing the lung coefficient, wall area of the bronchioles and Periodic Acid Schiff (PAS)-positive cells in the lung tissue. The CS-induced inflammation and oxidative stress was alleviated by soyasaponin I through reversing the levels of inflammatory cytokines and oxidative stress indicators. In addition, the phosphorylation of p38, JNK and ERK1/2 was activated in COPD model, and was reverted by soyasaponin I in the lung tissue. Collectively, the present study confirmed that soyasaponin I is an effective compound that attenuates the lung injury through inhibiting inflammatory response and oxidative stress via the mitogen-activated protein kinase (MAPK) signaling pathway.

This study aims to clarify the disruption of gut barrier and dysbiosis of the microbiota in an experimental macaque model with 6-year diabetes mellitus (DM), and provide evidence for the application of therapeutic strategies targeting the human microbiota in the future. A single intravenous injection of high-dose streptozotocin was used to induce the type 1 diabetes (T1D) macaque model. Hematoxylin-Eosin (HE) and Periodic Acid Schiff (PAS) staining were conducted to observe colon morphological changes. The composition of gut microbiota was detected using 16S rRNA gene sequencing, and bioinformatics analysis was adopted to predict alterations in the microbial phenotype and function. Obvious intestinal inflammation and decreased goblet cells were observed in T1D macaques. 16S rRNA gene sequencing suggested a significantly different β diversity of the microbiota in the T1D group, where expanded Proteobacteria (dominantly Escherichia-Shigella) and Actinomycetota (formerly known as Actinobacteria) replaced the dominance of Bacillota (formerly known as Firmicutes) and Bacteroidota (formerly known as Bacteroidetes), indicating an imbalance in the microbial composition. Archaea was identified as a biomarker between groups. Moreover, with the reduction of beneficial bacteria (Lactobacillaceae) and the increase of pro-inflammatory bacteria and opportunistic pathogens (Enterobacteriaceae), the phenotypes of the microbiota were reversed, resulting in abnormal up- (e.g., carbohydrate and amino acid metabolism) or down-regulation (e.g., protein digestion and absorption) of multiple metabolic pathways. There were intestinal structural disorders and gut microbiota dysbiosis in T1D macaques, indicating that strategies targeting gut microbiota may be effective to treat metabolic diseases like DM.

Uncontrolled activation of autophagy following ischemia/reperfusion (I/R) injury leads to cell death. The superfamily of ankyrin repeat proteins (N-Ank protein) was reported to be involved in autophagy regulation and cardiac protection. Bioinformatics analysis was performed (GSE61592 and GSE160516) and ten N-Ank proteins were differentially expressed in I/R models. Retinoic acid-induced protein 14 (RAI14), a member of N-Ank protein family, was upregulated in I/R-injured cardiac tissue and was first selected for research. A mouse I/R model was established by ligating the left anterior descending coronary artery to induce 90 min of ischemia, followed by 72 h of reperfusion. RAI14 was found upregulated in ischemic penumbra. RAI14 overexpression in cardiac tissue by injecting adeno-associated virus-9-RAI14 plasmid system via tail vein improved cardiac function and reduced infarct and apoptosis. Furthermore, the activated autophagy in ischemic penumbra of I/R mice was reversed by RAI14 overexpression along with decreased microtubule-associated protein 1 light chain 3 beta (LC3) II and increased autophagy receptor p62 expressions. RAI14 silence showed an opposite effect. A cell model was established by using mouse cardiomyocytes HL-1 underwent hypoxia/reoxygenation (H/R) treatment. Similarly, H/R also enhanced RAI14 expression and RAI14 overexpression inhibited H/R-induced apoptosis and autophagy in HL-1 cells. Mechanistically, autophagy inhibitor, the AKT/mTOR pathway, was found to be suppressed in mouse and cell models whereas RAI14 overexpression activated this pathway. Collectively, we demonstrated that compensatory increase of RAI14 inhibited I/R-induced myocardial injury by preventing excessive autophagy through activating the AKT/mTOR pathway, which providing an idea to explore strategies for preventing I/R injury.

The ratio of the second and fourth digits (2D:4D) is a morphological marker reflecting fetal exposure to sex steroid hormones. This ratio exhibits sexual dimorphism, with males typically showing a significantly lower ratio than females, which results from higher androgen exposure during the fetal period. While studies in humans have suggested a relationship between sexual orientation and the 2D:4D ratio, this relationship in rodents remains elusive. Here, we investigated this relationship using rats as an experimental model. We found that male rats exhibited significantly shorter 2D length than females, resulting in a lower 2D:4D ratio in males, similar to humans. Observations of sexual behavior revealed that males that ejaculated during the first mating test exhibited shorter 2D length compared to males those that did not ejaculate. When males were classified into two groups based on 2D length (long-2D and short-2D groups), short-2D males were more sexually active than long-2D males. Additionally, only short-2D males showed a preference for female odors. These findings suggest that, in rats, 2D length is a useful morphological marker reflecting sexual activity and preference. Furthermore, they provide evidence supporting the potential use of the 2D:4D ratio as a tool for studying the relationship between sexual orientation and the 2D:4D ratio in humans.

Translation regulation is crucial for cellular homeostasis. Recent studies have demonstrated that, in addition to the conventional AUG start codon, eukaryotic mRNA can also possess non-canonical start codons. These non-canonical start codons, including non-AUG codons, can be found both upstream and downstream of the conventional AUG start codon. Translation of these non-canonical open reading frames (ORFs) has been implicated in the development of diseases, such as cardiac diseases, neurodegeneration and cancer development. Non-AUG translation initiation is regulated by eukaryotic initiation factor (eIF) 2A and eIF2D; however, their target non-canonical ORFs, roles in disease development, and the underlying precise mechanisms of translation regulation remain poorly understood. To address these gaps, we generated mice lacking either or both of Eif2a and Eif2d genes on an ICR background and investigated their cardiac function using echocardiography. The results indicated that simultaneous disruption of both Eif2a and Eif2d led to perinatal cardiac impairment, as evidenced by a significant reduction in cardiac contractility as measured by ejection fraction. Furthermore, the absence of phenotypic changes in single knockouts of either Eif2a or Eif2d suggests that eIF2A and eIF2D function redundantly in their molecular roles. These findings underscore the importance of non-AUG translation initiation in maintaining cardiac function and suggest its broader implications in other physiological and pathological processes. We propose the Eif2a and Eif2d double-knockout mice as a novel stress-sensitive animal model to investigate the molecular mechanisms of translation regulation and their contribution to disease pathogenesis.