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The Wnt signaling pathway plays an important role in maintaining liver homeostasis and liver regeneration. In healthy livers， the Wnt signaling pathway is mostly inactive， but it is continuously overactivated during cell renewal or regeneration processes， as well as in certain pathological conditions， diseases， precancerous states， and cancers. Persistent liver cell injury often leads to chronic liver diseases such as liver fibrosis， liver cirrhosis， and liver cancer. This article summarizes the basic structural features of the Wnt signaling pathway and analyzes its important role in the pathological progression of various liver diseases， so as to provide new ideas for the prevention and treatment of liver diseases in clinical practice.
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Objective:To study the effects of intervention of Jiawei Tongmai Huazhi Decoction in the Wnt/β-catenin pathway on apoptosis of lesion cells in mice with adenomyosis (AM); To discuss its mechanism of action.Methods:The AM mouse model was established using tamoxifen. The mice were divided into model group, Jiawei Tongmai Huazhi Decoction group, and progesterone group according to random number table method, with 7 mice in each group. Additionally, a blank group of 7 female mice was set up. Jiawei Tongmai Huazhi Decoction group received oral administration of Jiawei Tongmai Huazhi Decoction at a dosage of 36.51 g/kg/day, once daily. The progesterone group received oral administration of progesterone at a dose of 0.32 mg/kg twice a week. The blank group and model group received oral administration of the same volume of physiological saline once daily. After 2 months of intervention, the morphology of uterine tissues was observed by HE staining. The levels of carbohydrate antigen 125 (CA125) and prolactin (PRL) in the serum were measured by ELISA. The mRNA levels of Wnt3a and β-catenin in uterine tissues were determined by PCR. The protein expressions of Wnt3a, β-catenin, Bax, and Bcl-2 in uterine tissues were detected by Western blot.Results:Compared with the model group, the levels of serum CA125 and PRL were reduced in the Jiawei Tongmai Huazhi Decoction group ( P<0.05). The protein expressions of Wnt3a, β-catenin, and Bcl-2 were also reduced ( P<0.05), the protein expressions of Wnt3a, β-catenin, and Bcl-2 decreased ( P<0.05), while the protein expressions of Bax increased ( P<0.05). Conclusion:Jiawei Tongmai Huazhi Decoction alleviates the progression of lesions by reducing serum CA125 and PRL levels in AM model mice, and can down-regulate Bcl-2 expression and up-regulate Bax expression, promoting apoptosis of ectopic lesion cells in mice. Its mechanism of action may be related to the inhibition of Wnt/β-catenin pathway related expression proteins.
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Objective To investigate the targeted differentiation ability of mouse bone marrow derived mesenchymal stem cells(BM-MSCs)and adipose-derived mesenchymal stem cells(AD-MSCs).Methods BM-MSCs and AD-MSCs were isolated and cultured from bone marrow of femur and white adipose tissue of groin of C57BL/6J mice respectively,and the two types of cells were induced by osteogenic,chondrogenic and adipogenic differentiation medium respectively.Alizarin red,alcian blue and oil red O staining were used to detect the differentiated degree of osteogenic,chondrogenic and lipogenic differentiation.Real-time fluorescence quantitative PCR(qPCR)was used to identify MSCs and detected expression levels of directed differentiation-related genes Runx2,Sp7(osteoblast),Sox9,Col2a1(chondroblast),Pparg and Cebpa(lipogenesis)to determine the directed differentiation ability of cells.Based on gene expression profiles of mouse and human BM-MSCs and AD-MSCs in GEO database GSE43804 and GSE122778,the differentially expressed genes and their enrichment signal pathways were analyzed.Results The cell morphology of BM-MSCs and AD-MSCs obtained by isolation and culture was different,and spindle-shaped morphology was more obvious in AD-MSCs.Both cells expressed CD29,CD44 and CD90,but did not express CD34 and CD45.AD-MSCs showed higher osteogenic and lipogenic differentiation than those of BM-MSCs after directed induction,while chondrogenic differentiation was lower in AD-MSCs than that of BM-MSCs(P＜0.05).After directional induction,expression levels of Runx2,Pparg and Cebpa mRNA were higher in AD-MSCs than those in BM-MSCs,and Sox9 mRNA expression levels were lower than those in BM-MSCs(P＜0.05).Highly expressed genes of AD-MSCs in mice and human were enriched in PPAR and WNT signaling pathways.Highly expressed genes of BM-MSCs were enriched in cartilage and bone developmental signaling pathways.Conclusion The osteogenic and adipogenic differentiation ability of mouse AD-MSCs is stronger than those of BM-MSCs,while the chondrogenic differentiation ability AD-MSCs is weaker than that of BM-MSCs.The activation status of PPAR,WNT,cartilages and skeletal system development signaling pathways plays an important regulatory role in determining the different directional differentiation potential of AD-MSCs and BM-MSCs.
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BACKGROUND:Wnt signaling pathway is overexpressed in degenerative intervertebral discs,and inhibition of its expression can delay the process of intervertebral disc degeneration.Therefore,Wnt signaling pathway is closely related to intervertebral disc degeneration. OBJECTIVE:To summarize the relationship between Wnt signaling pathway and intervertebral disc,especially the specific role and influence of Wnt signaling pathway in intervertebral disc degeneration. METHODS:The first author took"intervertebral disc,Wnt,cell proliferation,cell senescence,cell apoptosis,extracellular matrix"as the English search terms.PubMed,Web of science and OVID LWWSpringerlink were searched for articles published from 2000 to January 2023,and articles related to Wnt signaling pathway and disc degeneration were consulted.Totally 54 articles were reviewed by reading,collating and preserving. RESULTS AND CONCLUSION:(1)During intervertebral disc formation in embryos,Wnt signaling pathway is overexpressed,which is involved in intervertebral disc formation and promotes posterior extension of notochord.(2)During intervertebral disc degeneration,Wnt signaling pathway can inhibit cell proliferation by stagnating cell cycle,increase the expression of age-related proteins and promote cell aging by participating in oxidative stress,and participate in cell apoptosis by regulation of long non-coding RNA.(3)Wnt signaling pathway can also decrease extracellular matrix related protein synthesis,promote extracellular matrix degradation and accelerate intervertebral disc degeneration.(4)Wnt signaling pathway can promote cell regeneration by activating as early intervertebral disc formation signal and participate in inducing stem cells to differentiate into intervertebral disc cells to repair damaged intervertebral disc.For example,Wnt signaling pathway can induce stem cells from cartilage endplate cells to migrate and transform to intervertebral disc.
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BACKGROUND:Autoimmune regulator gene(Aire)and Wnt signaling pathway play an important role in the maintenance and differentiation of mouse embryonic stem cell pluripotency.However,whether the Wnt signal and Aire are involved in the differentiation of embryonic stem cells to thymic epithelial progenitor cells remains poorly understood. OBJECTIVE:To investigate the relationship of the Wnt signaling pathway and Aire with the differentiation of embryonic stem cells. METHODS:A two-step differentiation method was used to induce mouse embryonic stem cells to differentiate into endoderm and then into thymic epithelial progenitor cells.Mouse embryonic stem cells were infected with Aire shRNA lentivirus,and monoclonal stable strains were screened by puromycin.Mouse embryonic stem cells were collected on days 0,3 and 10 of the directed induction of differentiation after the induced differentiation by the two-step differentiation method.Cellular immunofluorescence,flow cytometry,western blot assay,and real-time qPCR were used to detect the expression changes of related genes and proteins. RESULTS AND CONCLUSION:(1)Immunofluorescence staining showed positive expression of SSEA1 and OCT4 on day 0 of targeted induction of differentiation.(2)Immunofluorescence staining showed double-positive expression of SOX17 and FOXA2 on day 3 of targeted induction of differentiation.(3)Flow cytometry results showed positive expression of EPCAM1,K5 and K8 on day 10 of targeted induction of differentiation.(4)Compared with undifferentiated mouse embryonic stem cells,the expressions of Wnt7a,β-catenin,and Gsk-3β proteins were elevated,and the expression level of Aire protein was decreased in induced differentiated thymic epithelial progenitor cells.(5)Compared with undifferentiated mouse embryonic stem cells,the expressions of Wnt7a,β-catenin,Gsk-3β and Aire mRNA were elevated in thymic epithelial progenitor cells.(6)Compared with normal cultured mouse embryonic stem cells and their ultimately differentiated thymic epithelial progenitor cells,the expression levels of Wnt7a,β-catenin and Gsk-3β proteins were reduced in mouse embryonic stem cells with knockdown of Aire genes and their final differentiated thymic epithelial progenitor cells.In conclusion,the Wnt signaling pathway and Aire are jointly involved in the process of targeted induction of differentiation of mouse embryonic stem cells into mouse thymic epithelial progenitor cells.
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ObjectiveTo explore the effect of Qingre Huayu Jianpi prescription （QHJ） on colitis-associated colorectal cancer （CAC） in mice， and its related mechanism. MethodC57BL/6 mice were randomly divided into four groups including the normal， model， QHJ low-dose （QHJ-L， 10 g·kg-1）， and QHJ high-dose （QHJ-H， 40 g·kg-1） groups. Azoxymethane （AOM） and dextran sodium sulfate （DSS） were combined to chemically build a CAC mouse model for 14 weeks. Each drug group was given intragastrically from the 5th week to the 14th week， once per day. An equal volume of water was fed to the normal and model groups. The mouse survival rate， colon length， weight， and pathological alterations were assessed. The protein expressions of Wnt-3a protein signaling （Wnt3a）， β-catenin， Non-phosphor-β-catenin （Non-p-β-catenin）， and cholesterol-binding glycoproteins 133 （CD133） were detected by Western blot. The localization and expression of the cluster of differentiation （CD） 80 and CD11 antigen-like family member B （CD11b） were detected by immunohistochemistry （IHC）. The colon organoids derived from CAC mice were isolated and cultured to detect the expression of Wnt signaling pathway-related proteins. ResultThe survival rate of the CAC mice was improved by QHJ treatment and the number of colon tumors was inhibited significantly. Compared with those in the normal group， the expression levels of Wnt3a， β-catenin， Non-p-β-catenin， and CD133 in colon tissues in the model group were significantly increased （P<0.05， P<0.01）. Compared with those in the model group， the levels of Wnt3a and β-catenin in the QHJ-L group were significantly decreased （P<0.01）， and the protein levels of Wnt3a， β-catenin， Non-p-β-catenin， and CD133 in the QHJ-H group were significantly decreased （P<0.05， P<0.01）. Meanwhile， the expression level of CD11b in the model group was significantly increased compared with that in the normal group while the CD80 level was significantly decreased （P<0.05， P<0.01）. Compared with those in the model group， CD11b in QHJ-L and QHJ-H groups was significantly decreased， and CD80 was significantly increased（P<0.05， P<0.01）. The expressions of Non-p-β-catenin and CD133 in colonic organoids of CAC model mice were significantly increased， while QHJ treatment could inhibit the expressions of Non-p-β-catenin and CD133 in colonic organoids （P<0.01）. ConclusionQHJ could inhibit the inflammation-cancer development in CAC mice， the mechanism of which might be related to regulating the microenvironment and inhibiting the over-activation of Wnt signaling.
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Diabetic osteoporosis （DOP） is a kind of bone complication caused by diabetes， which is characterized by the decrease of bone mineral density， the change of bone microstructure and the increase of bone fragility. The process of DOP is closely related to high glucose， insulin resistance， oxidative stress and other mechanisms. The Wnt/β-catenin signaling pathway plays an important role in mediating insulin resistance and bone metabolic balance in diabetes. Regulation of Wnt signal transduction promotes the expression of glycogen synthase kinase-3β（GSK-3β）phosphorylation and improves glucose and lipid metabolism. The Wnt/β-catenin signaling pathway is also an important way regulating osteocyte-driven bone remodeling， which not only plays an important regulatory role in the balance between osteoblasts and osteoclasts and improve bone metabolic homeostasis， but also promotes the expression of osteopontin， osteocalcin and type Ⅰ collagen， and improves bone proliferation and osteogenic differentiation by regulating the Wnt pathway. In recent years， the research of traditional Chinese medicine （TCM） in the prevention and treatment of DOP has gradually increased， and the exploration of TCM to interfere with the Wnt pathway to improve DOP has made some progress. This paper collects and summarizes the studies on the Wnt signaling pathway in glucose metabolism， bone metabolism and DOP worldwide in the past decade， as well as the related literature on the intervention of DOP by TCM compounds （classical and other compounds）， single Chinese medicine and TCM monomers based on the Wnt pathway， in order to provide a reference and direction for the development of new drugs for clinical prevention and treatment of DOP.
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Purpose: Pre-eclampsia (PE) is a pregnancy-related complication. Eucommia is effective in the treatment of hypertensive disorders in pregnancy, but the specific effects and possible mechanisms of Eucommia granules (EG) in PE remain unknown. The aim of this study was to investigate the effects and possible mechanisms of EG in PE rats. Methods: Pregnant Sprague Dawley rats were divided into five groups (n = 6): the control group, the model group, the low-dose group, the medium-dose group, and the high-dose group of EG. The PE model was established by subcutaneous injection of levonitroarginine methyl ester. Saline was given to the blank and model groups, and the Eucommia granules were given by gavage to the remaining groups. Blood pressure and urinary protein were detected. The body length and weight of the pups and the weight of the placenta were recorded. Superoxide dismutase (SOD) activity and levels of malondialdehyde (MDA), placental growth factor (PIGF), and soluble vascular endothelial growth factor receptor-1 (sFIt-1) were measured in the placenta. Pathological changes were observed by hematoxylin-eosin staining. Wnt/ß-catenin pathway-related protein expression was detected using Western blot. Results: Compared with the model group, the PE rats treated with EG had lower blood pressure and urinary protein. The length and weight of the pups and placental weight were increased. Inflammation and necrosis in the placental tissue was improved. SOD level increased, MDA content and sFIt-1/PIGF ratio decreased, and Wnt/ß-catenin pathway-related protein expression level increased. Moreover, the results of EG on PE rats increased with higher doses of EG. Conclusions: EG may activate the Wnt/ß-catenin pathway and inhibit oxidative stress, inflammation, and vascular endothelial injury in PE rats, thereby improving the perinatal prognosis of preeclamptic rats. EG may inhibit oxidative stress, inflammation, and vascular endothelial injury through activation of the Wnt/ß-catenin pathway in preeclampsia rats, thereby improving perinatal outcomes in PE rats.
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Animais , Ratos , Pré-Eclâmpsia , Estresse Oxidativo , Via de Sinalização Wnt , Inflamação , Animais de LaboratórioRESUMO
OBJECTIVE@#To investigate the mechanism of drug reversing resistance of Agaricus blazei extract FA-2-b-β on T cell acute lymphoblastic leukemia (T-ALL) cell lines.@*METHODS@#Cell proliferation was detected by CCK-8 assay; the apoptosis, cell cycle mitochondrial membrane potential, and intracellular rhodamine accumulation were detected by flow cytometry, and apoptosis-related gene and protein expression were detected by qPCR and Western blot; the membrane surface protein MDR1 was observed by immunofluorescence microscopy.@*RESULTS@#Different concentrations of FA-2-b-β significantly inhibited proliferation and induced apoptosis of CCRF-CEM and CEM/C1 (P<0.05), and CCRF-CEM cell cycle were arrested at S phase, and CEM/C1 cells were arrested at G0/G1 phase. Western blot and qPCR results show that FA-2-b-β inhibited ABCB1、ABCG2、CTNNB、MYC and BCL-2 expression, but upregulated Bax expression. In addition, FA-2-b-β reversed the resistance characteristics of CEM/C1 drug-resistance cells, which decreased mitochondrial membrane potential, and significantly increased the intracellular rhodamine accumulation, and weakening of the expression of the membrane surface protein MDR1. With the Wnt/β-catenin inhibitor (ICG001), the process was further intensified.@*CONCLUSION@#Agaricus Blazei Extract FA-2-b-β inhibits cell proliferation, promotes apoptosis, regulates the cell cycle, reduces mitochondrial energy supply, and down-regulate MDR1 expression to reverse the resistance of CEM/C1, which all suggest it is through regulating the Wnt signaling pathway in T-ALL.
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Humanos , Leucemia-Linfoma Linfoblástico de Células T Precursoras , Via de Sinalização Wnt , Leucemia-Linfoma Linfoblástico de Células Precursoras/metabolismo , Apoptose , Resistência a Múltiplos Medicamentos , Proteínas de Membrana , Linhagem Celular Tumoral , Proliferação de CélulasRESUMO
As a rare and heterogeneous group of non-Hodgkin lymphomas, most of peripheral T-cell lymphomas (PTCL) are highly aggressive and usually do not respond well to chemotherapy. The research found that the disorder of canonical Wnt signaling pathway is involved in the pathogenesis of some subtypes of PTCL. Canonical Wnt signaling pathway plays an important role in the development and differentiation of T-lymphocytes and tumorigenesis. In recent years, exploring the relationship between canonical Wnt signaling pathway and the development of PTCL is expected to provide new ideas and targets for the treatment of PTCL.
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Objective:To investigate the mechanism of Xianglian Huazhuo Decoction in treating chronic atrophic gastritis (CAG) rats by detecting the protein and gene expressions of Wnt1, GSK-3β, β-catenin in Wnt signaling pathway.Methods:Totally 60 Wistar rats were randomly divided into blank group and model group. The CAG model was established by MNNG free drink combined with sodium salicylate gavage combined with abnormal hunger and satiety. After model establishment, the rats were randomly divided into model group, Xianglian Huazhuo Group and folic acid group. Folic acid group received the folic acid water solution 1.3 mg/kg for gavage; the Xianglian Huazhuo Group received Xianglian Huazhuo Decoction 15.5 g/kg for gavage; the blank group and the model group received the same volume of purified water for gavage. After 8 weeks of intervention, the general state and the atrophic degree of gastric mucosa were observed. The expressions of Wnt1, GSK-3β and β-catenin were detected by immunohistochemical method, and the mRNA expression of β-catenin expression was detected by real-time quantitative polymerase chain reaction PCR.Results:The general condition of rats in Xianglian Huazhuo group was improved, the atrophy of gastric mucosa was improved, and the effect was better than that in folic acid group. Compared with the model group, the positive area ratio of Wnt1 decreased in Xianglian Huazhuo group ( P<0.05), and the positive area ratio of β-catenin decreased in folic acid group and Xianglian Huazhuo group ( P<0.05 or P<0.01), the positive area ratio of GSK-3β increased ( P<0.05), and the β-catenin mRNA level of Xianglian Huazhuo group decreased ( P<0.05). Conciusion Xianglian Huazhuo Decoction may increase GSK-3β by downregulating abnormal expression of Wnt1, promoting β-catenin to degrade normally, thereby blocking the abnormal activation of the Wnt signaling pathway, delaying or reversing CAG carcinogenesis to a certain extent, and exerting therapeutic effects.
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Objective:To observe the repair effect and possible mechanism of Dipsacus saponins Ⅵ on tibial fracture model rats.Methods:Thirty Sprague Dawley (SD) rats were randomly divided into model group, intervention group, and combination group, with 10 rats in each group, to establish a tibial fracture rat model using the sawing method. On the second day after surgery, the intervention group was intraperitoneally injected with 10 mg/kg of Chuanduduan saponin Ⅵ; The combination group received intraperitoneal injection of Dipsacus saponins Ⅵ 10 mg/kg and XAV939 1 mg/animal; The model group was intraperitoneally injected with 0.2 ml of physiological saline solution and 0.2 ml of dimethylsulfoxide (DMSO) solution; Once a day, continuous intervention for 14 days. After 2 to 4 weeks of intervention, Micro CT scan and X-ray scan were used to observe the fracture healing status; After 4 weeks of intervention, the wet weight of the tibia was detected; Hematoxylin eosin (HE) staining was used to observe the pathological changes of callus tissue; The Western blot method was used to detect the expression level of callus tissue β- catenin (β-catenin), p-β-catenin, glycogen synthase kinase 3β (GSK-3 β) and Runt related transcription factor 2 (Runx2) protein.Results:After 2 and 4 weeks of intervention, the bone volume fraction (BV/TV), number of trabeculae (Tb.N), Lane Sandhu score, and callus volume in the intervention group were higher than those in the model group (all P<0.05); After 2 and 4 weeks of intervention, the BV/TV, Tb.N, Lane Sandhu score, and callus volume in the combined group were lower than those in the intervention group (all P<0.05). The wet weight of the tibia in the intervention group was higher than that in the model group at 4 weeks after intervention ( P<0.05); The wet weight of the tibia in the combined group was lower than that in the intervention group ( P<0.05). The HE staining results showed that the model group had fibrous tissue hyperplasia and more bone trabeculae, but the maturity was not high and the thickening was not significant; The intervention group formed more bony callus, with orderly arrangement of bone trabeculae, partially mature, and obvious mineralization, consistent with the direction of stress; The combined group formed more cartilaginous and fibrous callus, with more mineralization at the edge of the cartilaginous callus and the formation of bone trabeculae. Abundant capillaries can be observed in the gaps. The expression level of Runx2 and p-β-catenin/β-catenin protein in callus tissue of the intervention group was higher than that of the model group, the protein expression GSK-3 β level was lower than that of the model group (all P<0.05); The expression level of Runx2 and p-β-catenin/β-catenin protein in the callus tissue of the combined group was lower than that of the intervention group; the protein expression level of GSK-3β was higher than that of the intervention group (all P<0.05). Conclusions:Dipsacus saponins Ⅵ can effectively promote fracture repair in tibial fracture model rats; It is possible to plays a role by activating the Wnt/β-catenin signaling pathway.
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Objective: To investigate the therapeutic effects of Tuina (Chinese therapeutic massage) in a knee osteoarthritis (KOA) rat model and its influence on proteins associated with the Wnt/β-catenin signaling pathway. Methods: A total of 32 specific-pathogen-free grade Sprague-Dawley rats were used. Eight rats were randomly selected as the control group (CG). The remaining 24 rats underwent intra-articular injections with 0.2 mL of 4％ papain to prepare the KOA rat models. After the model was established, the 24 rats were randomly and equally assigned to 3 groups, including a model group (MG), a Tuina group (TG), and a positive medicine group (PMG), with 8 rats in each group. The Lequesne score was applied to evaluate the success of model development. After the model was successfully established, the CG did not receive any intervention, and the TG was treated with local, clockwise annular Rou-Kneading around the knee joint with the thumbs. The pressure in the longitudinal direction was 3 N, and the frequency was designed to be 120-140 times/min for 15 min, followed by flexing the joint 10 times. The PMG was intragastrically administered with celecoxib [24 mg/(kg·bw)] every day. These interventions were performed once a day, 6 d per week, for a total of 4 weeks. After treatment, the Lequesne score was applied again to assess the severity of the KOA in the rats; hematoxylin-eosin (HE) staining and a mixture of equal volumes of aqueous solutions of safranin O-fast green were used to stain and observe the cartilage morphology and structure; the modified Mankin score was applied to evaluate the pathology; enzyme-linked immunosorbent assay method was used to quantify the C-telopeptide fragments of type Ⅱ collagen (CTX-Ⅱ) and cartilage oligomeric matrix protein (COMP); Western blotting was then applied to quantify Wnt4, β-catenin, matrix metalloproteinase 13 (MMP-13), and bone morphogenetic protein 2 (BMP-2) protein expression; immunohistochemistry was conducted to determine the percentage of collagen type X (ColX)-positive cells. Results: The Lequesne score of the TG and PMG was both lower than that of the MG (P<0.01); the HE staining, safranin O-fast green stained morphology and structure, and modified Mankin scores of the TG and the PMG were also better than those in the MG (P<0.01). Compared with the CG, the amounts of CTX-Ⅱ and COMP in the serum were significantly increased (P<0.01); the expression of Wnt4, β-catenin, MMP-13, and BMP-2 proteins in the cartilage tissue was significantly increased (P<0.01), and the percentage of ColX-positive chondrocytes was significantly increased (P<0.01) in the MG. In comparison with those in the MG, the amounts of CTX-Ⅱ and COMP were significantly decreased (P<0.01), the expression of Wnt4, β-catenin, MMP-13, and BMP-2 proteins was significantly decreased (P<0.01), and the percentage of ColX-positive chondrocytes was significantly decreased (P<0.01) in the TG and PMG. Compared with the PMG, the contents of CTX-Ⅱ and COMP and the expression of Wnt4, β-catenin, MMP-13, and BMP-2 proteins were decreased (P<0.05 or P<0.01); the percentage of ColX-positive chondrocytes was significantly decreased (P<0.01) in the TG. Conclusion: Tuina can relieve the degeneration of KOA, and the mechanism may be related to the down-regulation of the Wnt/β-catenin signaling pathway, the decrease in MMP-13 and BMP-2 protein expression, the reduction in chondrocyte extracellular matrix degradation, and slowing down the terminal cell differentiation.
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Objective @#To explore the regulatory mechanism of ginsenoside Rb1 on focal cerebral ischemia-reperfusion injury ( CIRI) ．@*Methods @# A total of 60 C57 / BL mice were randomly divided into 6 groups (n = 10) : shamoperated group ，CIRI model group ，ginsenoside Rb1 low －，medium －，and high-dose group and nimodipine (positive control) group．The surgical method was used to construct the focal CIRI mouse model．The neurological function scores and behavioral tests were performed，and Nissl staining was utilized to detect the number of nissl bodies in the hippocampus．The effect of ginsenoside Rb1 on the molecule expression of the Wnt signaling pathway in the hippocampus was detected by qPCR , Western blot and immunohistochemistry assays．The regulatory mechanism of ginsenoside Rb1 was investigated through molecular docking and co-precipitation assays. @*Results @#Compared with the CIRI model group，the addition of ginsenoside Rb1 reduced the neurological function scores of mice (P＜0. 05) ，shortened the time passing the balance beam (P＜0. 05) ，but increased the time entering the correct arm (P＜0. 05) and the swinging time and climbing time of mice (P ＜0. 05 ) ，indicating that ginsenoside Rb1 could effectively resume the function of the nervous system in mice and improve the behavioral ability of model mice．After ginsenoside Rb1 treatment，axis inhibition protein 2 (Axin2) and glycogen synthase kinase-3 β ( GSK- 3 β) in the hippocampus decreased，whereas the expression of Wnt3a，Wnt1 and β-catenin increased．@*Conclusion@#The ginsenoside Rb1 can improve neurological function of the CIRI mouse model and increase the number of Nissl bodies in the hippocampus，which is correlated with the activation of the Wnt signaling pathway，and it may be neuroprotective against focal CIRI during stroke treatment.
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Objective To investigate the effect of transcription factor specific protein5(Sp5)silencing on Wnt signaling pathway correlated factors and cell proliferation ability in mouse embryo palatal mesenchymal cells(mEPMCs).Methods mEPMCs of 14.5 d pregnant C57BL/6J mice were isolated and cultured in vitro.Cell source was identified by immunofluorescence staining.Lentivirus transfection technique was used to silence the expression of Sp5 gene in mEPMCs,and the transfection efficiency was verified by Western blot assay.Follow-up experiments were set up with the blank control group,the no-load virus group and the slience group(the Sp5-shRNA group).The protein and mRNA expression levels of β-catenin,GSK-3β,Wnt3a and CyclinD1 were detected by Western blot assay and RT-qPCR after transfection for 72 h in each group.Cell proliferation capacity was detected by CCK-8.The proliferation rate of 5-Ethynyl-2'-deoxyuridine(EdU)positive cells was detected by immunofluorescence assay.Cell cycle was detected by flow cytometry.Results mEPMCs were successfully isolated,and Sp5 expression was silenced.Western blot and RT-qPCR results showed that the protein and mRNA expressions of β-catenin,GSK-3β,Wnt3a and CyclinD1 were significantly higher in the Sp5-shRNA group than those in the blank control group and the no-loaded virus group(P<0.05).The proliferative ability and the proliferative rate of EdU positive cells were higher in the Sp5-shRNA group than those in the blank control group and the no-loaded virus group(P<0.05).The proportion of mEPMCs in S phase was higher in the Sp5-shRNA group than that in the blank control group and the no-loaded virus group(P<0.05).Conclusion Sp5 in silenced mEPMCs can participate in palate development and promote the proliferation of mEPMCs by regulating Wnt signaling pathway.
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Objective:To observe the inhibitory effects of Huangqi Jiedu Decoction on lung metastasis of breast cancer in nude mice; To explore the mechanism of intervening epithelial mesenchymal transformation (EMT) induced by Wnt/β-catenin signaling pathway.Methods:Totally 30 nude mice were divided into model group, adriamycin group and Huangqi Jiedu Decoction low-, medium-, and high-dosage groups according to random number table method. Each group was injected subcutaneously with mouse breast cancer 4T1 cells to construct tumor - bearing nude mice model. Huangqi Jiedu Decoction low-, medium- and high-dosage groups were intragastrically administrated with Huangqi Jiedu Decoction 17.82, 35.64 and 71.28 g/kg; adriamycin group was injected intraperitoneally adriamycin 0.05 g/kg; model group was intragastrically administrated with normal saline of the same volume for 21 d. Tumor volume was measured at 9, 15, and 21 days after modeling. After the end of administration, the tumor tissue was separated, the tumor weight was measured, and the tumor inhibition rate was calculated. The lung tissue was Isolated,, the number of lung metastatic nodules and the inhibition rate of lung metastasis was counted. HE staining was used to observe the tissue morphology and evaluate the effectiveness of the model. The protein expressions of β-catenin, E-Cadherin and Vimentin in lung tissue were detected by Western Blot. The mRNA levels of β-catenin, E-Cadherin and Vimentin in lung tissue were detected by real-time fluorescent quantitative PCR.Results:Compared with the model group, the tumor volume and mass of Huangqi Jiedu Decoction low-, medium- and high-dosage groups decreased ( P<0.01); the number of pulmonary metastasis nodules in Huangqi Jiedu Decoction high-dosage group significantly decreased ( P<0.01); the mRNA and protein expressions of β-catenin and Vimentinm decreased in the Huangqi Jiedu Decoction low-, medium- and high-dosage groups ( P<0.01), and the protein and mRNA expressions of E-Cadherin increased in the Huangqi Jiedu Decoction high-dosage group ( P<0.01). Conclusion:Huangqi Jiedu Decoction can effectively inhibit the growth and lung metastasis of breast cancer transplanted tumor, and the mechanism may be to down-regulate the expression of key molecules in the Wnt/β-catanin signaling pathway, thereby inhibiting the EMT process, so as to inhibit the lung metastasis of breast cancer.
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Objective: To investigate the expression and significance of protease activated receptor 2 (PAR2) in ovarian epithelial carcinoma. Methods: PAR2 mRNA expression levels in 410 cases of epithelial ovarian carcinoma and 88 cases of human normal ovary were analyzed from cancer Genome Atlas (TCGA) database and tissue genotypic expression database (GTEx). Immunohistochemical (IHC) staining of PAR2 protein was performed in 149 patients with ovarian cancer who underwent primary surgical treatment at Cancer Hospital of Chinese Academy of Medical Sciences. Then the relationship between mRNA/protein expression of PAR2 and clinicopathological features and prognosis was analyzed. Gene functions and related signaling pathways involved in PAR2 were studied by enrichment analysis. Results: The mRNA expression of PAR2 in epithelial ovarian carcinoma was significantly higher than that in normal ovarian tissue (3.05±0.72 vs. 0.33±0.16, P=0.004). There were 77 cases showing positive and 19 showing strong positive of PAR2 IHC staining among the 149 patients, accounting for 64.4% in total. PAR2 mRNA/protein expression was closely correlated with tumor reduction effect and initial therapeutic effect (P<0.05). Survival analysis showed that the progression free survival time (P=0.033) and overall survival time (P=0.011) in the group with high PAR2 mRNA expression was significantly lower than that in the low PAR2 mRNA group. Multivariate analysis showed tumor reduction effect, initial therapeutic effect were independent prognostic factors on both progression-free survival and overall survival (P<0.05). The progression-free survival (P=0.016) and overall survival (P=0.038) of the PAR2 protein high expression group was significantly lower than that of the low group. Multivariate analysis showed PAR2 expression, initial treatment effect and chemotherapy resistance were independent prognostic factors on both progression-free survival and overall survival (P<0.05). Based on Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG), PAR2 target genes were mainly enriched in function related to intercellular connection, accounting for 40%. Gene enrichment analysis (GSEA) showed that the Wnt/β-catenin signaling pathway (P=0.023), the MAPK signaling pathway (P=0.029) and glycolysis related pathway (P=0.018) were enriched in ovarian cancer patients with high PAR2 mRNA expression. Conclusions: PAR2 expression is closely related to tumor reduction effect, initial treatment effect and survival of ovarian cancer patients. PAR2 may be involved in Wnt/β-catenin signaling pathway and intercellular connection promoting ovarian cancer invasion and metastasis.
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Feminino , Humanos , Carcinoma Epitelial do Ovário , Receptor PAR-2 , Neoplasias Ovarianas/patologia , Prognóstico , RNA Mensageiro/metabolismoRESUMO
Objective To investigate the effect of microRNA (miR)-138 regulation of Wnt signaling pathway on the biological behavior of human glioma cells in vitro. Methods Glioma cell lines U-87MG and U251 were selected and randomly divided into blank group, miR-NC group, miR-138 mimics group and miR-138 inhibitor group. Real-time PCR was used to detect the miR-138 expression in each group; MTT, flow cytometry, Transwell assay and scratch assay were used to detect proliferation, apoptosis, invasion and migration ability of each group respectively, and Western blotting was used to detect Wnt pathway-related protein expression in each group. Results The miR-138 expression level was higher in the miR-138 mimics group compared with the remaining 3 groups, and that in the miR-138 inhibitor group was lower than that in the blank group and the miR-NC group (P<0. 05) ; Compared with the blank group, the cell proliferation rate was lower in the miR-138 mimics group and higher in the miR-138 inhibitor group, and was time-dependent (P<0. 05) ; The apoptosis rate in the miR-138 mimics group was higher than that in the blank group, miR-NC group, and miR-138 inhibitor group, while the apoptosis rate in the miR-138 inhibitor group was lower than that in the rest other groups (P<0. 05) ; The number of cell-invading cells in the miR-138 mimics group was lower than that in the blank group, miR-NC group, and miR-138 inhibitor group, while all miR-138 inhibitor group were higher than the remaining three groups (P<0. 05) ; The cell migration rate of miR-138 mimics group was lower than that of blank group, miR-NC group and miR-138 inhibitor group, while all miR-138 inhibitor group were higher than the remaining three groups (P<0. 05) ; Wnt3a, Wntl, glycogen synthase kinase 3(3(GSK-3(3) and (3-catenin protein expression in the miR-138 mimics group was lower than that in the blank group, miR-NC group, and miR-138 inhibitor group; While miR-138 inhibitor groups were higher than the remaining three groups(P<0. 05). Conclusion MiR-138 overexpression effectively inhibite the proliferation, invasion and migration of glioma cells and promote their apoptosis, probably achieved by pathway inhibition of the Wnt signaling pathway.
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SUMMARY OBJECTIVE: Celiac disease is an autoimmune disease characterized by an abnormal immune response occurring in the small intestine linked to consumption of food containing gluten in individuals with a genetic predisposition. Dysregulation of Wnt signal transduction plays a role in the pathogenesis of many diseases including autoimmune diseases like celiac disease. In this study, the correlation of Wnt pathway gene expressions with each other and the correlation with clinical data were researched in pediatric celiac disease cases grouped according to the Marsh classification. METHODS: Gene expression levels of FZD8, DVL2, LRP5, RHOA, CCND2, CXADR, and NFATC1, which are involved in the Wnt pathway, were determined using quantitative real-time polymerase chain reaction in 40 celiac disease and 30 healthy individuals. RESULTS: All cases with the short height symptom were observed to be in Marsh 3b/3c groups (p=0.03). The gene expressions of DVL2, CCND2, and NFATC1 were high in the Marsh 3b group, and these genes showed positive correlation with each other (p=0.002). LRP5 and CXADR gene expressions were lower in the Marsh 3b group compared to other Marsh groups, and these genes showed a positive correlation with each other (p=0.003). CCND2 gene expression was associated with Marsh 3b group, diarrhea, and vomiting symptoms. DVL2 gene expression was correlated with Marsh 2 group and constipation symptom (p<0.05). CONCLUSION: Wnt signaling in the early stages of the disease of Marsh 1-2 involves high expression of LRP5 and CXADR genes, while expression of these two genes reduces, and DVL2, CCND2, and NFATC1 gene expressions clearly increase with a transduction variation observed from Marsh 3a stage when villous atrophy begins to form. It appears that the Wnt pathway may contribute to disease progression through expression changes.
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Abstract Tooth development depends on a series of reciprocal signaling interactions between the oral epithelium and ectomesenchyme. This study aimed to investigate the role of CK14, a protein involved in Wnt-1/β-catenin signaling, in odontogenesis and the development of odontomas. This cross-sectional, retrospective, immunohistochemical study analyzed 30 compound odontomas, 30 complex odontomas, and 17 tooth germs. Higher immunoexpression of CK14 was observed in odontogenic epithelial cells of tooth germs (p < 0.001) and odontogenic epithelial cells of odontomas (p < 0.001). There was higher immunoexpression of Wnt-1 and β-catenin proteins in epithelial cells of tooth germs (p = 0.002 and p < 0.001, respectively), as well as in the ectomesenchyme of odontomas (p = 0.003 and p < 0.001, respectively). β-Catenin was moderately and significantly correlated with CK14 in the membrane of reduced enamel epithelial cells in odontomas (p = 0.007). Higher immunoexpression of CK14 was observed in the odontogenic epithelium during the bud and cap stages and lower immunoexpression in the internal enamel epithelium during the bell stage. In odontomas, lower expression of Wnt-1/β-catenin and higher immunoexpression of CK14 were found in odontogenic epithelial cells, especially adjacent to the mineralized material resembling the tooth formed in these lesions.
Resumo O desenvolvimento dentário depende de uma série de interações de sinalização recíproca entre o epitélio oral e o ectomesênquima. O objetivo deste estudo foi investigar o papel da CK14 das vias WNT-1/β-catenina na odontogênese e no desenvolvimento de odontomas. Este estudo transversal, retrospectivo, imuno-histoquímico analisou 30 odontomas compostos, 30 odontomas complexos e 17 germes dentários. A CK14 apresentou maior imunoexpressão em células epiteliais odontogênicas de germes dentários (p<0,001) e em células epiteliais odontogênicas de odontomas (p<0,001). A Wnt-1 e a β-catenina apresentaram maior imunoexpressão de proteínas nas células epiteliais dos germes dentários (p = 0,002 e p<0,001, respectivamente), bem como no ectomesênquima dos odontomas (p = 0,003 e p < 0,001, respectivamente). A β-catenina correlacionou-se moderada e significativamente com a CK14 na membrana de células epiteliais reduzidas do esmalte em odontomas (p = 0,007). Maior imunoexpressão da CK14 foi observada no epitélio odontogênico nos estágios de botão e capuz com menor imunoexpressão no epitélio interno do órgão do esmalte no estágio de sino. Nos odontomas, foi observado menor expressão de Wnt-1/β-catenina e maior imunoexpressão da CK14 presente nas células epiteliais odontogênicas, especialmente, vizinhas ao material mineralizado semelhante ao dente formado nessas lesões.