This study investigates the therapeutic effect and underlying mechanism of Compound Ziyin Granules(CZYG) on postmenopausal osteoporosis(PMOP) induced by bilateral ovariectomy in rats. Six-month-old female SD rats were randomly divided into sham-operated(sham) group, ovariectomy(OVX) model group, high-, medium-, and low-dose CZYG groups, and alendronate sodium(AS) group. After 30 days of model establishment, treatment was administered by gavage once daily for 8 weeks, followed by sample collection. Enzyme-linked immunosorbent assay(ELISA) was used to measure serum levels of calcium ions, alkaline phosphatase(AKP), estrogen(E_2), osteoprotegerin(OPG), osteocalcin(BGP), tartrate-resistant acid phosphatase(TRAP), and type Ⅰ procollagen N-terminal propeptide(PINP). Hematoxylin-eosin(HE) staining was used to observe the histopathological changes in the femurs of rats, while micro-computed tomography(micro-CT) was used to analyze the microstructure of the distal femur. Western blot analysis was performed to measure the expression levels of bone metabolism-related proteins, including wingless-type MMTV integration site family member 2(Wnt2), β-catenin, low-density lipoprotein receptor-related protein 5(LRP5), glycogen synthase kinase-3β(GSK-3β). The mRNA expression levels of Wnt2, β-catenin, LRP5, GSK-3β, p-GSK-3β were determined by quantitative real-time PCR(qRT-PCR). Thirty days after bilateral ovariectomy, compared to the sham group, the OVX group showed significant increases in body weight and significant decreases in uterine coefficient. After 8 weeks of treatment, compared to the OVX group, CZYG(medium and high doses) and AS reduced body weight, with high-dose CZYG and AS significantly increasing the uterine coefficient. Serum levels of AKP and TRAP were significantly elevated, while levels of calcium, E_2, BGP, and OPG were significantly decreased in the OVX group. Compared to the OVX group, CZYG and AS significantly reduced serum levels of AKP and TRAP, while high-dose CZYG and AS notably increased the levels of E_2, BGP, OPG, and PINP. Micro-CT and HE staining results indicated that CZYG(medium and high doses) and AS significantly increased bone tissue volume, trabecular number, bone mineral density, and improved the microstructure of the femur. Compared to the OVX group, high-dose CZYG and AS significantly upregulated the protein and mRNA expression levels of Wnt2, β-catenin, and LRP5, and downregulated the phosphorylation level of p-GSK-3β. These results suggest that CZYG can improve PMOP by promoting estrogen secretion, improving bone metabolism indicators, increasing trabecular number and bone mineral density. Its mechanism may be related to the regulation of the Wnt/β-catenin signaling pathway.
Animals ; Female ; Rats, Sprague-Dawley ; Osteoporosis, Postmenopausal/genetics* ; Rats ; Wnt Signaling Pathway/drug effects* ; Humans ; Drugs, Chinese Herbal/administration & dosage* ; beta Catenin/genetics* ; Osteoprotegerin/metabolism* ; Ovariectomy ; Calcium/blood* ; Bone Density/drug effects*

Renal ischemia-reperfusion injury (IRI) is a major contributor to acute kidney injury (AKI), leading to substantial morbidity and mortality. Spexin (SPX), a 14-amino acid endogenous peptide involved in metabolic regulation and immune modulation, has not yet been studied in the context of chronic treatment and renal IRI. This study evaluated the effects of exogenous SPX on renal function, histopathological changes, and molecular pathways in both IRI-induced injured and healthy kidneys. Twenty-eight male BALB/c mice were divided into four groups: control, SPX, IRI, and SPX+IRI. IRI was induced by 30 minutes of bilateral renal ischemia followed by 6 hours of reperfusion. Renal injury markers, histopathological changes, inflammatory mediators, apoptotic markers, and fibrosis-related proteins were analyzed. SPX significantly exacerbated IRI-induced kidney injury by activating the Wnt/β-catenin signaling pathway and promoting the upregulation of pro-inflammatory, pro-apoptotic, and pro-fibrotic mediators. It is noteworthy that SPX exerted more severe deleterious nephrotoxic effects in the healthy kidney compared to those observed in the IRI-induced injured kidney. These findings indicate that chronic treatment with SPX administration may have intrinsic pro-inflammatory, pro-apoptotic and fibrotic properties, raising concerns about its therapeutic potential. Further research is needed to clarify its physiological role and therapeutic implications in kidney diseases.
Animals ; Reperfusion Injury/chemically induced* ; Male ; Mice, Inbred BALB C ; Mice ; Acute Kidney Injury/metabolism* ; Kidney/blood supply* ; Peptide Hormones/adverse effects* ; Apoptosis/drug effects* ; Wnt Signaling Pathway/drug effects* ; Disease Models, Animal

Metastasis remains the primary cause of cancer-related mortality worldwide. Circulating tumor cells (CTCs) represent critical targets for metastasis prevention and treatment. Traditional Chinese medicine may prevent lung cancer metastasis through long-term intervention in CTC activity. Tiao-Shen-Zhi-Ai Formular (TSZAF) represents a Chinese medicine compound prescription utilized clinically for lung cancer treatment. This study combined three principal active ingredients from TSZAF into a novel TSZAF monomer combination (TSZAF mc) to investigate its anti-metastatic effects and mechanisms. TSZAF mc demonstrated significant inhibition of proliferation, migration, and invasion in CTC-TJH-01 and LLC cells, while inducing cellular apoptosis in vitro. Moreover, TSZAF mc substantially inhibited LLC cell growth and metastasis in vivo. Mechanistically, TAZSF mc significantly suppressed the Wnt/β-catenin signaling pathway and CXCL5 expression in lung cancer cells and tissues. Additionally, TAZSF mc notably reduced neutrophil infiltration in metastatic lesions. These findings indicate that TSZAF mc inhibits lung cancer growth and metastasis by suppressing the Wnt/β-catenin signaling pathway and reducing CXCL5 secretion, thereby decreasing neutrophil recruitment and infiltration. TSZAF mc demonstrates potential as an effective therapeutic agent for lung cancer metastasis.
Lung Neoplasms/genetics* ; Wnt Signaling Pathway/drug effects* ; Animals ; Humans ; Drugs, Chinese Herbal/pharmacology* ; Mice ; Neoplasm Metastasis/prevention & control* ; Cell Proliferation/drug effects* ; Cell Line, Tumor ; Neutrophil Infiltration/drug effects* ; Down-Regulation/drug effects* ; Cell Movement/drug effects* ; beta Catenin/genetics* ; Apoptosis/drug effects* ; Mice, Inbred C57BL ; Male ; Neoplastic Cells, Circulating/drug effects*

Human cytomegalovirus (HCMV) poses a significant risk of neural damage during pregnancy. As the most prevalent intrauterine infectious agent in low- and middle-income countries, HCMV disrupts the development of neural stem cells, leading to fetal malformations and abnormal structural and physiological functions in the fetal brain. This review summarizes the current understanding of how HCMV infection dysregulates the Wnt signaling pathway to induce fetal malformations and discusses current management strategies.
Humans ; Cytomegalovirus Infections/virology* ; Wnt Signaling Pathway ; Pregnancy ; Female ; Cytomegalovirus/physiology* ; Pregnancy Complications, Infectious/virology* ; Congenital Abnormalities/virology* ; Animals

Objective To investigate the effects of folate deficiency on changes in histone H3 lysine 4 (H3K4) mono-methylation (me1)-marked enhancers and the molecular mechanism underpinning the folate deficiency-induced neural tube defects (NTD). Methods Mouse embryonic stem cells (mESCs) were cultured in the folate-free DMEM medium (folate-deficient group) and the DMEM medium containing 4 mg/L folate (normal control group),respectively.Chromatin immunoprecipitation sequencing (ChIP-seq) was performed for H3K4me1. The mouse model of folate-induced NTD was established,and transcriptome sequencing (RNA-seq) was performed for the brain tissue of fetal mice to reveal the differential expression profiles.The results were validated through real-time quantitative polymerase chain reaction (RT-qPCR).The activity of the differential peak regions of H3K4me1 was verified through the luciferase reporter assay. Results The folate content in the mESCs cultured in the folate-free medium reduced compared with that in the normal control group (P=0.008).The H3K4me1-maked enhancers in the mESCs cultured in the folate-free medium induced significant changes in intronic regions,and these changes were concentrated in metabolic and energy metabolism processes (q=9.56×10^-48,P=1.28×10^-47).The differentially expressed genes harboring H3K4me1-marked enhancers in mESCs were mainly enriched in the Wnt signaling pathway (q=0.004,P=0.004 7).ChIP-qPCR results confirmed that H3K4me1 binding decreased in the differential peak regions of the Ldlrap1 gene (P=0.008),Camta1 gene (P=0.002),and Apc2 gene (P=0.012).The H3K4 demethylase inhibitor T-448 effectively reversed the H3K4me1 binding in the differential peak regions of the aforementioned genes (P=0.01).The results of RNA-seq for the brain tissue of NTD fetal mice showed significant enrichment of the differentially expressed genes in the Wnt signaling pathway (P=1.52×10^-5).The enrichment of differential peak regions of H3K4me1-marked enhancers in Apc2,Ldlrap1,and Camta1 genes in the brain tissue also showed significant changes.The differential peak region in Apc2 exhibited transcription factor activity (P=0.020). Conclusion Folate deficiency may affect changes in H3K4me1-marked enhancers to participate in the regulation of neural tube closure genes,thereby inducing the occurrence of NTD.
Neural Tube Defects/genetics* ; Animals ; Mice ; Folic Acid Deficiency/complications* ; Histones/metabolism* ; Folic Acid/metabolism* ; Methylation ; Mouse Embryonic Stem Cells/metabolism* ; Wnt Signaling Pathway ; Lysine/metabolism* ; Chromatin Immunoprecipitation Sequencing

Objective To investigate the role and mechanism of RNA-Specific adenosine deaminase 3 (Adar3) in regulating macrophage polarization during Coxsackievirus B3(CVB3)-induced viral myocarditis (VM). Methods Bone marrow-derived macrophages (BMDM) from mice were cultured in vitro and induced into M1/M2 macrophages using interferon-gamma (IFN-γ)/lipopolysaccharide (LPS) or interleukin 4 (IL-4), respectively. The mRNA expression levels of Adar1, Adar2, and Adar3 in each group of cells were assessed by real-time quantitative PCR (qRT-PCR). Specific siRNAs targeting the Adar3 gene were designed, synthesized, and transiently transfected into M2 macrophages. The mRNA levels of M2 polarization-related marker genes-including arginase 1 (Arg1), chitinase 3-like molecule 3 (YM1/Chi3l3), and resistin-like molecule alpha (RELMα/FIZZ1)-were detected by qRT-PCR. RNA sequencing was performed to analyze the signaling pathways affected by Adar3. The expression levels of Wnt/β-catenin signaling pathway were further validated using qRT-PCR and Western blot. The adeno-associated virus overexpressing Adar3 was designed, synthesized, and injected into mice via tail vein. Three weeks later, a myocarditis mouse model was established. After an additional week, the phenotype and function of cardiac macrophages, as well as multiple indicators of VM (including echocardiography, body weight, histopathology and serology) were examined. Additionally, the protein levels of the Wnt/β-catenin signaling pathway were assessed. Results Compared to M0-type macrophages, the expression level of Adar3 was significantly increased in M2-type macrophages. After transfection of Adar3 siRNA, the mRNA levels of Arg1, YM1 and FIZZ1 in M2 macrophages were downregulated. RNA sequencing revealed 149 upregulated genes and 349 downregulated genes. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis and subsequent validation experiments indicated that Adar3 modulated the Wnt/β-catenin signaling pathway. In vivo experiments demonstrated that Adar3 overexpression alleviated the cardiac dysfunction of VM mice. The proportion of M1 macrophages in the heart decreased, while the proportion of M2 macrophages increased. At the same time, the Adar3 overexpression activated the Wnt/β-catenin signaling pathway. Conclusion Adar3 promotes macrophage polarization toward the M2 phenotype by activating the Wnt/β-catenin signaling pathway, thereby alleviating VM.
Animals ; Adenosine Deaminase/metabolism* ; Macrophages/immunology* ; Wnt Signaling Pathway/genetics* ; Myocarditis/immunology* ; Mice ; Coxsackievirus Infections/metabolism* ; Male ; Mice, Inbred BALB C ; Enterovirus B, Human/physiology* ; beta Catenin/genetics*

OBJECTIVES: To investigate the role and mechanism of Brahma-related gene 1 (Brg1) in regulating the Wnt/β-catenin signaling pathway in a bronchopulmonary dysplasia (BPD) model. METHODS: Wild-type C57BL/6 and Brg1^f1/f1 mice were randomly divided into four groups: wild-type control, wild-type BPD, Brg1^f1/f1 control, and Brg1^f1/f1 BPD (n=5 each). Immortalized mouse pulmonary alveolar type 2 cells (imPAC2) were cultured, and Brg1 gene was knocked down using lentivirus transfection technology. Cells were divided into three groups: control, empty vector, and Brg1 knockdown. Hematoxylin and eosin staining and immunofluorescence were used to detect pathological changes in mouse lung tissue. Western blot and real-time fluorescent quantitative PCR were used to measure Brg1 protein and mRNA expression levels in mouse lung tissue. Western blot and immunofluorescence were used to detect the expression of homeodomain-containing protein homeobox (HOPX), surfactant protein C (SPC), and Wnt/β-catenin signaling pathway proteins in mouse lung tissue and imPAC2 cells. The CCK8 assay was used to assess the proliferation of imPAC2 cells, and co-immunoprecipitation was performed to verify the interaction between Brg1 and β-catenin proteins in imPAC2 cells. RESULTS: Compared to the Brg1^f1/f1 control group and wild-type BPD group, the Brg1^f1/f1 BPD group showed increased alveolar diameter and SPC protein expression, and decreased relative density of pulmonary vasculature and HOPX protein expression (P<0.05). Compared to the control group, the Brg1 knockdown group showed increased cell proliferation ability, protein expression levels of SPC, Wnt5a and β-catenin, and β-catenin protein fluorescence intensity, along with decreased HOPX protein expression (P<0.05). An interaction between Brg1 and β-catenin proteins was confirmed. CONCLUSIONS The Brg1 gene may promote the proliferation of alveolar type 2 epithelial cells by regulating the Wnt/β-catenin signaling pathway, thus influencing the occurrence and development of BPD.
Animals ; DNA Helicases/genetics* ; Transcription Factors/genetics* ; Wnt Signaling Pathway/physiology* ; Nuclear Proteins/genetics* ; Mice ; Bronchopulmonary Dysplasia/etiology* ; Mice, Inbred C57BL ; beta Catenin/physiology* ; Disease Models, Animal ; Cell Proliferation ; Lung/pathology* ; Male

OBJECTIVE: To investigate the effect of Arsenic trioxide (ATO) combined with Norcantharidin (NCTD) on the proliferation and apoptosis of Jurkat cells, and to evaluate its effect on the proliferation and apoptosis of acute T-lymphoblastic leukemia (T-ALL) based on the Wnt/β-catenin signaling pathway. METHODS: Jurkat cell lines were used as the study subjects and treated with different concentrations of ATO (0, 2, 4, 8, 16 μmol/L) and NCTD (0, 10, 25, 50, 100 μmol/L) for 72 hours, and the cell proliferation was detected by CCK-8. Meanwhile, flow cytometry was used to detect the apoptosis rate, EdU staining to detect cell proliferation viability, cell clone formation assay to assess cell cloning ability, Transwell assay to assess cell invasion ability, and Western blot to detect apoptosis and the expression of Wnt/β-catenin signaling pathway-related proteins. RESULTS: Compared with the control group, both ATO and NCTD effectively inhibited Jurkat cell proliferation when used alone, and the inhibition effect was more significant when used in combination ( P < 0.05). The combination significantly increased the apoptosis rate of Jurkat cells ( P < 0.05). Meanwhile, the combination significantly decreased the proliferation vitality and clone formation ability of the cells ( P < 0.05), and inhibited the invasion ability of Jurkat cells ( P < 0.05). Western blot analysis showed that the combination of ATO and NCTD significantly up-regulated the expression of pro-apoptotic proteins Bax and E-cadherin, and down-regulated the expression of anti-apoptotic proteins Bcl-2, c-myc and Cyclin D1 ( P < 0.05). CONCLUSION The combination of ATO and NCTD had a synergistic effect in inhibiting proliferation and promoting apoptosis in Jurkat cells, which may be related to the inhibition of Wnt/β-catenin signaling pathway.
Humans ; Apoptosis/drug effects* ; Jurkat Cells ; Cell Proliferation/drug effects* ; Arsenic Trioxide ; Wnt Signaling Pathway/drug effects* ; Bridged Bicyclo Compounds, Heterocyclic/pharmacology* ; beta Catenin/metabolism* ; Arsenicals/pharmacology* ; Oxides/pharmacology*

Excessive generation of reactive oxygen species (ROS) can lead to oxidative-antioxidative imbalance in the organism, resulting in oxidative stress. Hematopoietic stem/progenitor cells (HSPCs) exhibit high sensitivity to changes in ROS levels, and high levels of ROS can impair self-renewal capacity of HSPCs, leading to oxidative damage and even death. Wnt/β-catenin signaling pathway regulates hematopoiesis and plays an important role in determining the fate of stem cells, such as self-renewal, proliferation and differentiation of HSPCs. Studies have shown that Wnt/β-catenin signaling pathway is also closely related to oxidative stress. This article summarizes the relevant literature, and reviews the role of Wnt/β-catenin signaling pathway in oxidative stress, its impact on hematopoietic system, and the current research status of related mechanisms.
Oxidative Stress ; Humans ; Wnt Signaling Pathway ; Hematopoietic Stem Cells ; Reactive Oxygen Species/metabolism* ; Hematopoietic System/metabolism* ; beta Catenin/metabolism* ; Hematopoiesis

OBJECTIVE: To investigate the Wnt signaling pathway and miRNAs mechanism of extracts of Plastrum Testudinis (PT) in the treatment of osteoporosis (OP). METHODS: Thirty female Sprague Dawley rats were randomly divided into 5 groups by random number table method, including sham group, ovariectomized group (OVX), ovariectomized groups treated with high-, medium-, and low-dose PT (160, 80, 40 mg/kg per day, respectively), with 6 rats in each group. Except for the sham group, the other rats underwent bilateral ovariectomy to simulate OP and received PT by oral gavage for 10 consecutive weeks. After treatment, bone mineral density was measured by dual-energy X-ray absorptiometry; bone microstructure was analyzed by micro-computed tomography and hematoxylin and eosin staining; and the expressions of osteogenic differentiation-related factors were detected by immunochemistry, Western blot, and quantitative polymerase chain reaction. In addition, Dickkopf-1 (Dkk-1) was used to inhibit the Wnt signaling pathway in bone marrow mesenchymal stem cells (BMSCs) and miRNA overexpression was used to evaluate the effect of miR-214 on the osteogenic differentiation of BMSCs. Subsequently, PT extract was used to rescue the effects of Dkk-1 and miR-214, and its impacts on the osteogenic differentiation-related factors of BMSCs were evaluated. RESULTS: PT-M and PT-L significantly reduced the weight gain in OVX rats (P<0.05). PT also regulated the bone mass and bone microarchitecture of the femur in OVX rats, and increased the expressions of bone formation-related factors including alkaline phosphatase, bone morphogenetic protein type 2, collagen type I alpha 1, and runt-related transcription factor 2 when compared with the OVX group (P<0.05 or P<0.01). Meanwhile, different doses of PT significantly rescued the inhibition of Wnt signaling pathway-related factors in OVX rats, and increased the mRNA or protein expressions of Wnt3a, β-catenin, glycogen synthase kinase-3β, and low-density lipoprotein receptor-related protein 5 (P<0.05 or P<0.01). PT stimulated the osteogenic differentiation of BMSCs inhibited by Dkk-1 and activated the Wnt signaling pathway. In addition, the expression of miR-214 was decreased in OVX rats (P<0.01), and it was negatively correlated with the osteogenic differentiation of BMSCs (P<0.01). MiR-214 mimic inhibited Wnt signaling pathway in BMSCs (P<0.05 or P<0.01). Conversely, PT effectively counteracted the effect of miR-214 mimic, thereby activating the Wnt signaling pathway and stimulating osteogenic differentiation in BMSCs (P<0.05 or P<0.01). CONCLUSION PT stimulates bone formation in OVX rats through β-catenin-mediated Wnt signaling pathway, which may be related to inhibiting miR-214 in BMSCs.
Animals ; MicroRNAs/genetics* ; Female ; Rats, Sprague-Dawley ; Wnt Signaling Pathway/genetics* ; Osteogenesis/genetics* ; Mesenchymal Stem Cells/cytology* ; Cell Differentiation/drug effects* ; Bone Density/drug effects* ; Ovariectomy ; Osteoporosis/drug therapy* ; beta Catenin/metabolism* ; Rats ; Intercellular Signaling Peptides and Proteins/metabolism* ; Drugs, Chinese Herbal/pharmacology*