ObjectiveTo evaluate the safety/efficacy of compound Kushen injection （CKI） by zebrafish model and explore the possible mechanism. MethodsZebrafish were exposed to different concentrations of CKI solution， and the mortality rate after 24 h was calculated. After exposure to sublethal concentration （10） and safe dose concentration （0） for 24 h， the safety of CKI was evaluated based on acridine orange staining for the liver， liver area changes， and liver histological sections. The inflammatory bowel disease （IBD） model of zebrafish was established with 2，4，6-trinitrobenzenesulfonic acid sol （TNBS）. The efficacy of CKI in the treatment of IBD was evaluated by the changes in the area of neutral red staining， the number of neutrophils， the area of alcian blue staining， and the contents of tight junction protein （Occludin）， zonula occludens-1 （ZO-1）， tumor necrosis factor（TNF）-α， and prostaglandin E₂（PGE₂） in the intestine of zebrafish. The mechanism of CKI in the treatment of IBD was predicted by network pharmacology and verified by real-time polymerase chain reaction（Real-time PCR）. ResultsIn the safety evaluation， compared with the blank group， the sublethal concentration of CKI could cause liver cell apoptosis， liver area reduction， loose and disordered arrangement of liver cell structure， obvious vacuolation， and other pathological characteristics of zebrafish， while these indicators showed no significant changes under safe dose conditions. In the effectiveness evaluation， compared with the model group， the safe dose of CKI could increase the neutral red staining area of the intestine in a dose-dependent manner and improve the intestinal phagocytosis function. It could reduce the accumulation of intestinal neutrophils， increase the alcian blue staining area， enhance intestinal goblet cell secretion， improve the contents of Occludin and ZO-1， and decrease the contents of TNF-α and PGE₂. Network pharmacology analysis identified 288 potential targets for CKI treatment of IBD. The top five targets obtained by protein-protein interaction （PPI） network analysis were epidermal growth factor receptor A（EGFRA）， protein kinase B1（Akt1）， heat shock protein 90（HSP90AA1）， homologous oncogene of avian sarcoma virus（SRC）， and protooncogene （JUN）. Kyoto encyclopedia of genes and genomes（KEGG） results showed that CKI may be related to the Wnt signaling pathway and cholinergic synaptic pathway in the treatment of IBD， and Real-time PCR results verified the above pathways. ConclusionExcessive use of CKI can induce obvious liver injury in zebrafish， while the rational use of CKI does not cause obvious toxic reactions and can achieve a therapeutic effect on IBD by regulating the Wnt pathway.

Objective To investigate the effect and mechanism of Efferocytosis Relatived LncRNA (EFRL) on homocysteine-induced atherosclerosis in macrophage efferocytosis. Methods RAW264.7 cells were cultured in vitro, and the Control group (0 μmol/L Hcy) and Hcy intervention group (100 μmol/L Hcy) were set up. After GapmeR transfection of macrophages with Hcy intervention, EFRL knockdown negative control group (Hcy combined with LNA-NC) and EFRL knockdown group (Hcy combined with LNA-EFRL) were set up. High-throughput sequencing was applied for different expression of LncRNA MSTRG. 88917.16 (EFRL), UCSC was used to analyze its conservation, CPC and CPAT were used to analyze its ability to encode proteins, and GO and KEGG were used to analyze related biological functions. The localization of LncRNA EFRL in macrophages was analyzed by nucleoplasmic separation and RNA-FISH. Quantitative real-time PCR was used to detect the expression levels of LncRNA EFRL and its target gene SPAST in Hcy-treated macrophages. The apoptosis rate of Jurkat cells induced by UV was detected by flow cytometry. In vitro efferocytosis assay combined with immunofluorescence technique was used to analyze macrophage efferocytosis. ELISA was used to detect the levels of interleukin 1β(IL-1β) and IL-18. Results The new LncRNA MSTRG.88917.16 was identified and named EFRL(Efferocytosis Relatived LncRNA). UCSC, CPC and CPAT analyses showed that LncEFRL is highly conserved and does not have the ability to encode proteins. GO and KEGG analyses suggested that LncEFRL may be involved in macrophage efferocytosis. LncRNA EFRL was localized in the nucleus of macrophages as determined by nucleoplasmic separation and RNA-FISH. In comparison to the Control group, the expression levels of LncRNA EFRL and its target gene SPAST in the Hcy group were increased. In comparison to the Control group (0 min), the apoptosis rate of the experimental group (15, 30 min) Annexin V is more than 85%. Compared with Hcy combined with LNA-NC group, Hcy combined with LNA-EFRL group had enhanced macrophage efferocytosis and reduced levels of inflammatory factors. Compared with Hcy combined with LNA-NC group, the expression level of SPAST in Hcy combined with LNA-EFRL group was decreased. Conclusion Inhibition of EFRL expression can alleviate the process of Hcy inhibiting macrophage efferocytosis, and the mechanism is related to the regulation of the downstream target gene SPAST by EFRL.
RNA, Long Noncoding/physiology* ; Animals ; Homocysteine ; Mice ; Macrophages/drug effects* ; Humans ; RAW 264.7 Cells ; Atherosclerosis/chemically induced* ; Apoptosis/genetics* ; Phagocytosis/genetics* ; Jurkat Cells ; Interleukin-1beta/genetics* ; Efferocytosis

Mechanical pain is one of the most common causes of clinical pain, but there remains a lack of effective treatment for debilitating mechanical and chronic forms of neuropathic pain. Recently, neurotoxin GsMTx4, a selective mechanosensitive (MS) channel inhibitor, has been found to be effective, while the underlying mechanism remains elusive. Here, with multiple rodent pain models, we demonstrated that a GsMTx4-based 17-residue peptide, which we call P10581, was able to reduce mechanical hyperalgesia and neuropathic pain. The analgesic effects of P10581 can be as strong as morphine but is not toxic in animal models. The anti-hyperalgesic effect of the peptide was resistant to naloxone (an μ-opioid receptor antagonist) and showed no side effects of morphine, including tolerance, motor impairment, and conditioned place preference. Pharmacological inhibition of TRPV4 by P10581 in a heterogeneous expression system, combined with the use of Trpv4 knockout mice indicates that TRPV4 channels may act as the potential target for the analgesic effect of P10581. Our study identified a potential drug for curing mechanical pain and exposed its mechanism.

Hypercholesterolemia is a significant risk factor for the development of atherosclerosis. 2',3',5'-Tri-O-acetyl-N ⁶-(3-hydroxyphenyl) adenosine (IMM-H007), a novel AMPK agonist, has shown protective effects in metabolic diseases. However, its impact on cholesterol and triglyceride metabolism in hypercholesterolemia remains unclear. In this study, we aimed to elucidate the effects and specific mechanisms by which IMM-H007 regulates cholesterol and triglyceride metabolism. To achieve this goal, we used Apoe ^-/- and Ldlr ^-/- mice to establish a hypercholesterolemia/atherosclerosis model. Additionally, hepatocyte-specific Ampka1/2 knockout mice were subjected to a 5-week high-cholesterol diet to establish hypercholesterolemia, while atherosclerosis was induced via AAV-PCSK9 injection combined with a 16-week high-cholesterol diet. Our results demonstrated that IMM-H007 improved cholesterol and triglyceride metabolism in mice with hypercholesterolemia. Mechanistically, IMM-H007 modulated the AMPKα1/2-LDLR signaling pathway, increasing cholesterol uptake in the liver. Furthermore, IMM-H007 activated the AMPKα1-FXR pathway, promoting the conversion of hepatic cholesterol to bile acids. Additionally, IMM-H007 prevented hepatic steatosis by activating the AMPKα1/2-ATGL pathway. In conclusion, our study suggests that IMM-H007 is a promising therapeutic agent for improving hypercholesterolemia and atherosclerosis through the activation of AMPKα.

The anterior cingulate cortex (ACC) has recently been proposed as a key player in the representation of itch stimuli. However, to date, little is known about the contribution of specific ACC interneuron populations to itch processing. Using c-Fos immunolabeling and in vivo Ca²⁺ imaging, we reported that both histamine and chloroquine stimuli-induced acute itch caused a marked enhancement of vasoactive intestinal peptide (VIP)-expressing interneuron activity in the ACC. Behavioral data indicated that optogenetic and chemogenetic activation of these neurons reduced scratching responses related to histaminergic and non-histaminergic acute itch. Similar neural activity and modulatory role of these neurons were seen in mice with chronic itch induced by contact dermatitis. Together, this study highlights the importance of ACC VIP⁺ neurons in modulating itch-related affect and behavior, which may help us to develop novel mechanism-based strategies to treat refractory chronic itch in the clinic.
Animals ; Pruritus/physiopathology* ; Vasoactive Intestinal Peptide/metabolism* ; Interneurons/metabolism* ; Gyrus Cinguli/metabolism* ; Mice ; Male ; Mice, Inbred C57BL ; Histamine ; Chloroquine ; Optogenetics ; Mice, Transgenic

ObjectiveTo investigate the potential value of thyroid hormones in cerebrospinal fluid in predicting autism-like behaviors induced by hypothyroidism in pregnant rats. MethodsTwelve pregnant Wistar rats were randomly divided into a control group and a hypothyroidism group （hypothyroidism model group）. Offspring from both groups had serum and cerebrospinal fluid levels of thyroid-stimulating hormone （TSH）， total triiodothyronine （TT3）， total thyroxine （TT4）， free triiodothyronine （FT3）， and free thyroxine （FT4） levels in serum and cerebrospinal fluid. Ultrasonic vocalization tests were conducted on postnatal day 2 （P2）， day 7 （P7）， and day 14 （P14）， while behavioral tests using the three-box social interaction test were performed on day 21（P21）. ResultsCompared with the control group， free T4 （FT4） levels in the cerebrospinal fluid of the hypothyroidism group were significantly reduced during the developmental period （P0-P21； P2： P<0.05； P7： P<0.05； P14： P<0.01； P21： P<0.01）， with no statistical difference between the two groups only at P0 （P>0.05）. In the ultrasonic vocalization （USV） tests， the number and duration of USVs in offsprings from the hypothyroidism group were significantly reduced compared with those of the control group on P2， P7 and P14： for USV counts （P2： P<0.05； P7： P<0.001； P14： P<0.01）； for USV duration （P2： P<0.05； P7： P<0.001； P14： P<0.001）. In the three-box social tests， offsprings of the hypothyroidism group showed significantly reduced sniffing time with unfamiliar rats at P21 compared to the control group （all P<0.001）. The FT4 levels in cerebrospinal fluid had a significantly positive correlation with USV counts （P7： r=0.883， P<0.05； P14： r=0.902， P<0.05） and sniffing time with unfamiliar rats （r=0.814， P<0.01）. ConclusionMeasuring free T4 in cerebrospinal fluid can predict autism-like behaviors in offsprings of rats induced by hypothyroidism during pregnancy.

Rodent models play a crucial role in research on human periodontal diseases,providing key evidence for investigation into the pathological mechanisms of periodontal bone defects.Relevant research in the field involves gene expression,inflammatory regulation mechanisms,host-microbial interactions,as well as disease resolution and healing processes.Research methodology in the field falls under 2 categories-periodontal inflammation models and surgical defect models.The former simulates periodontal defects by inducing periodontal diseases,while the latter constructs clinically simulated periodontal defects through surgical removal of periodontal tissue.However,the currently available animal models of periodontitis face challenges in simultaneously capturing the disease complexity,tracking dynamic repair processes,and meeting translational needs.Herein,we reviewed and summarized the methods and characteristics of periodontal disease modeling in recent years.We proposed the establishment of a multimodal assessment framework integrating technologies such as spatial transcriptomics,single-cell sequencing,and in vivo fluorescence imaging,which may serve as a critical pathway for overcoming existing research challenges.

Objective:To explore the application value of PAX1/JAM3 methylation detection by cervical self-collected specimen in cervical cancer screening and the management of premenopausal and postmenopausal women.Method:This study is a single center cross-sectional study. From January 2023 to November 2023, cervical self-collected and physician-collected specimens at the colposcopy clinic were detected the PAX1/JAM3 methylation (PAX1 m/JAM3 m) testing. The consistency between self-collected and physician-collected specimens for PAX1 m/JAM3 m detection were compared based on histopathology. In addition, the clinical efficacy of methylation detection with high-risk human papillomavirus (hrHPV), liquid-based cytology (LBC), and their combination for cervical cancer screening were compared in the study. Results:A total of 301 women were recruited to undergo referral colposcopy examination, and statistical analysis was conducted on 272 women with pathological and diagnostic information. Among them, 102 cases (37.5%) were diagnosed as normal cervical tissue or chronic cervicitis, 72 cases (26.4%) were cervical intraepithelial neoplasia grade 1 (CIN1), 43 cases (15.8%) were CIN2, 29 cases (10.7%) were CIN3, and 26 cases (9.6%) were cervical cancer. According to the minimum quantity formula, they were divided into a consistency cohort of 81 participants and a validation cohort of 191 participants. The consistency between cervical self-collected and physician-collected specimens for detecting PAX1 m/JAM3 m. Results from spearman correlation analysis showed a positive correlation between the self-collected and physician-collected results of PAX1 m/JAM3 m detection, and the correlation coefficient R values are 0.858 ( P<0.001) and 0.828 ( P<0.001). The sensitivity and specificity of PAX1 m/JAM3 m detection for diagnosing CIN2 or more severe lesions (CIN2+) were 77.6% [95% confidence interval ( CI) 65.3%-86.4%] and 87.2% (95% CI 80.5%-91.9%), respectively. In clinical performance comparison, the sensitivity of PAX1 m/JAM3 m combined with HPV16/18 detection, 89.7% (95% CI 79.2%-95.2%), was the same as that of hrHPV detection in CIN2+and 96.0% (95% CI 80.4%-99.3%) in CIN3+, which is higher than 92.0% (95% CI 75.0%-97.8%) of hrHPV and 82.6% (95% CI 62.9%-93.0%) of LBC or the combination of sPAX1 m/JAM3 m and LBC low-grade and higher squamous intraepithelial lesion testing [87.0% (95% CI 67.9%-95.5%)]. Conclusions:Self-collected specimens by women for detection of PAX1 and JAM3 methylation as a promising screening tool for cervical cancer has operational and clinical feasibility. The methylation test can optimize the current cervical cancer screening plan, reduce the number of referral women with false positive diagnosis to colposcopy, and is of great significance for reducing fertility protection and preventing missed diagnosis in women of childbearing age.

Breast cancer is the most common cancer in women and one of the deadliest cancers worldwide.According to the distribution of tumor tissue,breast cancer can be divided into invasive and non-invasive forms.The cancer cells in invasive breast cancer pass through the breast and through the immune system or systemic circulation to different parts of the body,forming metastatic breast cancer.Drug resistance and distant metastasis are the main causes of death from breast cancer.Research on breast cancer has attracted extensive attention from researchers.In vitro construction of tumor models by tissue engineering methods is a common tool for studying cancer mechanisms and anticancer drug screening.The tumor microenvironment consists of cancer cells and various types of stromal cells,including fibroblasts,endothelial cells,mesenchymal cells,and immune cells embedded in the extracellular matrix.The extracellular matrix contains fibrin proteins(such as types Ⅰ,Ⅱ,Ⅲ,Ⅳ,Ⅵ,and Ⅹcollagen and elastin)and glycoproteins(such as proteoglycan,laminin,and fibronectin),which are involved in cell signaling and binding of growth factors.The current traditional two-dimensional(2D)tumor models are limited by the growth environment and often cannot accurately reproduce the heterogeneity and complexity of tumor tissues in vivo.Therefore,in recent years,research on three-dimensional(3D)tumor models has gradually increased,especially 3D bioprinting models with high precision and repeatability.Compared with a 2D model,the 3D environment can better simulate the complex extracellular matrix components and structures in the tumor microenvironment.Three-dimensional models are often used as a bridge between 2D cellular level experiments and animal experiments.Acellular matrix,gelatin,sodium alginate,and other natural materials are widely used in the construction of tumor models because of their excellent biocompatibility and non-immune rejection.Here,we review various natural scaffold materials and construction methods involved in 3D tissue-engineered tumor models,as a reference for research in the field of breast cancer.

During long-term electrocardiogram (ECG) monitoring, various types of noise inevitably become mixed with the signal, potentially hindering doctors' ability to accurately assess and interpret patient data. Therefore, evaluating the quality of ECG signals before conducting analysis and diagnosis is crucial. This paper addresses the limitations of existing ECG signal quality assessment methods, particularly their insufficient focus on the 12-lead multi-scale correlation. We propose a novel ECG signal quality assessment method that integrates a convolutional neural network (CNN) with a squeeze and excitation residual network (SE-ResNet). This approach not only captures both local and global features of ECG time series but also emphasizes the spatial correlation among ECG signals. Testing on a public dataset demonstrated that our method achieved an accuracy of 99.5%, sensitivity of 98.5%, and specificity of 99.6%. Compared with other methods, our technique significantly enhances the accuracy of ECG signal quality assessment by leveraging inter-lead correlation information, which is expected to advance the development of intelligent ECG monitoring and diagnostic technology.
Electrocardiography/methods* ; Humans ; Signal Processing, Computer-Assisted ; Neural Networks, Computer ; Algorithms