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OBJECTIVE To explore the mechanism of Naozhenning granules in regulating mitochondrial energy metabolism in hippocampal tissue of multiple cerebral concussion （MCC） model rats. METHODS SPF grade Wistar rats were used to prepare MCC models using the “free fall impact method”. The successfully modeled rats were divided into model group， piracetam group， and Naozhenning granule low-dose， medium-dose and high-dose groups， and a normal group was also set up， with 8 rats in each group. Rats in each treatment group orally administered corresponding drugs at doses of 0.324 g/kg for the piracetam group and 2.25， 4.5 and 9 g/kg for the Naozhenning granule low-dose， medium-dose and high-dose groups； the normal group and model group were given equal volumes of normal saline； once a day， for 14 consecutive days. The motor exploration ability， learning and memory ability of rats were tested； the adenosine triphosphate （ATP） content in the hippocampal tissue of rat was detected； the changes in the mitochondrial structure of hippocampal tissue was observed； the fluorescence intensity of mitochondrial dynamin- related protein 1 （Drp1）， mitochondrial fission protein 1 （Fis1）， mitochondrial fusion 1 （Mfn1）， and optic atrophy protein 1 （Opa1） were detected in the hippocampal tissue of rat； the protein expression levels of peroxisome proliferator activated receptor gamma coactivator-1α（PGC-1α）， nuclear respiratory factor-1（NRF-1），mitochondrial transcription factor A（TFAM）， Wnt-3a，β-catenin in hippocampal tissue of rat were detected. RESULTS Compared with the normal group， the total exercise distance， number of central grid entries， number of upright positions， new object recognition index， mitochondrial ATP content， fluorescence intensity of Mfn1 and Opa1， the protein expression levels of PGC-1α、NRF-1、TFAM、Wnt-3a、 β-catenin in the model group were significantly reduced （P＜0.01）， while the rest time and fluorescence intensity of Drp1 and Fis1 in hippocampal tissue were significantly increased （P＜0.01）. The results of transmission electron microscopy showed that the mitochondria in the hippocampal tissue were significantly swollen， with a large number of broken and reduced cristae， and some mitochondria had myeloid changes in the membrane. Compared with the model group， the levels/contents of the above indicators in rats of each administration group showed varying degrees of reversal， and most of the differences were statistically significant （P＜0.05 or P＜0.01）； the degree of mitochondrial swelling in the hippocampal tissue was reduced， with a small amount of broken and reduced cristae， fuzzy fractures appeared in local areas of the rough endoplasmic reticulum. CONCLUSIONS Naozhenning granules can improve the motor exploration， learning and memory abilities of MCC model rats， repair neuronal damage， and exert neuroprotective effects. Its mechanism may be related to activating Wnt/β-catenin signaling pathway，maintaining the balance of mitochondrial division and fusion，and promoting mitochondrial biosynthesis.
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ObjectiveThis study aims to investigate the mechanism in which Celastrus orbiculatus extract （COE） affects the proliferation and differentiation of gastric organoids and the expression of Lgr5 and thus reverses the precancerous lesions of gastric cancer （PLGC） by regulating the leucine-rich repeat containing G protein-coupled receptor 5 （Lgr5）/Wingless （Wnt）/β-catenin signaling pathway based on a gastric organoid injury model. MethodGastric organoids were established based on stem cells of the mouse gastric gland. Gastric organoid injury models were constructed by treating gastric organoids with N-methyl-N'-nitro-N-nitrosoguanidine （MNNG， 0.02 mg·L-1）. Gastric organoid injury models were randomly divided into normal group， model group （0.02 mg·L-1 MNNG）， low， medium， and high dose （5， 10， 20 mg·L-1） groups of COE， and Wnt inhibitor Dickkopf-related protein 1 （DKK1） （0.5 mg·L-1） group， and they were treated with respective agents for 24 h. The number and volume of gastric organoids under different drug concentrations were observed under a microscope. The viability of the gastric organoid injury models was detected by Methyl thiazolyl tetrazolium （MTT） assay. The morphology and pathology of gastric organoids were observed using Hematoxylin and Eosin （HE） staining. The expression levels of Lgr5， Mucin2 （MUC2）， Mucin5AC （MUC5AC）， Mucin6 （MUC6）， Wnt， and β-catenin in gastric organoids under different drug concentrations were detected by Western blot （WB）. ResultCompared with the normal group， the number， volume， and activity of gastric organoids in the model group were decreased （P<0.01）， while the expressions of Lgr5， MUC2， Wnt， and β-catenin were significantly increased （P<0.01）. The expressions of MUC5AC and MUC6 were significantly decreased （P<0.01）. Compared with the model group， the number and volume of gastric organoids in the low， medium， and high dose groups of COE were all improved （P<0.01）， and the vitality of gastric organoids was significantly enhanced （P<0.01）. The effect was the most significant at a COE concentration of 20 mg·L-1 （P<0.01）. The expressions of Lgr5 and MUC2 in the medium and high dose groups of COE were significantly reduced （P<0.01）， while the expression of MUC5AC and MUC6 were significantly increased in the low， medium， and high dose groups of COE （P<0.05， P<0.01）. Compared with the model group， Wnt inhibitors could promote the expression of MUC5AC and MUC6 in gastric organoids （P<0.05， P<0.01） and reduce the expression of MUC2， Wnt， and β-catenin. In addition， the combined use of COE at high concentrations and Wnt inhibitors could further promote this trend （P<0.01）. ConclusionCOE inhibits the Wnt/β-catenin pathway by inhibiting the expression of Lgr5， MUC2， Wnt， and β-catenin and promoting the expression of MUC5AC and MUC6， thus promoting the proliferation and differentiation of gastric organoids and reversing the PLGC process.
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Objective:To study the effects of Shuxuening injection(SXN)on Wnt/β-catenin signaling pathway in hippocampus of rats with cerebral ischemia.Methods:SD rats were divided into three groups:Sham operation group(Sham),middle cerebral artery occlusion group(MCAO)and MCAO+SXN treatment group(MCAO+SXN).The model of cerebral ischemia in rats was prepared by MCAO.The rats with cerebral ischemia were treated with SXN by caudal vein injection.Zea-Longa scoring criteria and balance beam test were employed to evaluated neurological function of rats.HE staining were used to observe the changes of inflammatory cells infiltration the hippocampal CAI region.The expression of β-catenin in hippocampal CA1 region was observed by immunofluorescence staining.The mRNA and pro-tein expressions of caspase-3,cyclooxygenase-2(COX-2)and endothelial nitric oxide synthetase(eNOS)in hippocam-pus were detected by real time RT-PCR and Western Blot,respectively.Results:SXN can SXN can improve the neuro-logical dysfunction of cerebral ischemia rats.The inflammatory cells infiltration in hippocampal CAI region was decreased,and the expression of β-catenin,caspase-3 and COX-2 was decreased,while the expression of eNOS was upregulated.Conclusion:SXN protects against cerebral ischemia by inhibiting Wnt/β-catenin signaling pathway against inflammation response,oxidative stress and apoptosis of nerve cells in rats.
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Objective:To explore the inhibitory mechanism of herb cake-partitioned moxibustion on tumor growth in colitis-associated colorectal cancer(CAC)based on histone lysine demethylase 4D(KDM4D). Methods:Inbred male Sprague-Dawley rats were randomly divided into a normal group,a CAC group,a herb cake-partitioned moxibustion group,and an inhibitor group.Except the normal group,rats in the other three groups were treated with azoxymethane(AOM)combined with dextran sulfate sodium(DSS)to make CAC rat models.Rats in the normal group and the CAC group did not receive interventions;rats in the herb cake-partitioned moxibustion group received moxibustion at Qihai(CV6)and bilateral Tianshu(ST25),2 cones for one point each time,once a day for 30 d with 1-day rest every week;rats in the inhibitor group received intraperitoneal injection of KDM4D inhibitor,5-chloro-8-hydroxyquinoline(5-c-8HQ),once a day for 30 d.After intervention,the general condition,colon length,tumor number and volume,and histopathological colon changes were observed.The expression of adenomatous polyposis coli(APC),axis inhibitor(Axin),cyclin D1,matrix metalloproteinase(MMP)-7 and MMP-9 mRNAs were detected by real-time quantitative polymerase chain reaction.The proliferating cell nuclear antigen(PCNA),cleaved caspase3,KDM4D,APC,and Axin proteins were detected by immunohistochemistry. Results:Compared with the normal group,the general condition was poor,the colon length was significantly shortened(P<0.01),the number and volume of colonic tumors were increased(P<0.01),the structure of glandular duct was obviously disordered with"back-to-back"and cowall phenomenon,and also high-grade adenocarcinoma formed;the protein expression levels of PCNA and KDM4D were significantly increased(P<0.01),while cleaved caspase3,APC,and Axin were significantly reduced(P<0.01);the mRNA expression levels of cyclin D1,MMP-7,and MMP-9 were significantly increased(P<0.01),while APC and Axin were significantly reduced(P<0.01)in the CAC group.Compared with the CAC group,the general condition was improved,the length of colon was significantly increased(P<0.01),the number and volume of the colonic tumors were reduced(P<0.05),and the colon tissues showed epithelial cell proliferation with enlarged and deep staining nuclei,dysplasia and inflammatory cell infiltration;the protein expression levels of PCNA and KDM4D were significantly reduced(P<0.01),while the cleaved caspase3,APC,and Axin were significantly increased(P<0.01);the mRNA expression levels of cyclin D1,MMP-7,and MMP-9 were reduced(P<0.05),while the APC and Axin were increased(P<0.05)in the colon tissues of rats in the herb cake-partitioned moxibustion group and the inhibitor group. Conclusion:Herb cake-partitioned moxibustion regulated abnormally expressed KDM4D in CAC rats,activated APC and Axin,the upstream molecules of Wnt/β-catenin pathway,inhibited abnormally activated downstream molecules of Wnt/β-catenin pathway.This may be a key mechanism of herb cake-partitioned moxibustion in inhibiting CAC tumor growth.
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Objective To investigate the effect and mechanism of Shuangzhu Kangxian Prescription(Astragali Radix,bran-fried Atractylodis Macrocephalae Rhizoma,vinegar-prepared Rhizoma Curcumae,Bupleuri Radix,Salviae Miltiorrhizae Radix et Rhizoma,Litchi Semen)on improving hepatitic fibrosis induced by carbon tetrachloride(CCl4)in rats based on the Wnt/β-catenin signaling pathway.Methods The rat model of hepatitic fibrosis was replicated by subcutaneous injection of 3.0 mL·kg-1 40%CCl4 twice a week for 8 weeks.SD rats were randomly divided into blank control group(n=8),model group(n=7),positive control group(n=7,intragastric administration of 43.19 mg·kg-1 silymarin),low-dose Chinese medicine group(n=6,intragastric administration of 4.3 g·kg-1Shuangzhu Kangxian Prescription),medium-dose Chinese medicine group(n=6,intragastric administration of 8.6 g·kg-1Shuangzhu Kangxian Prescription),high-dose Chinese medicine group(n=7,intragastric administration of 17.2 g·kg-1Shuangzhu Kangxian Prescription).The continuous intragastric administration was given once a day for 4 consective weeks,in addition to the blank control group,the other groups continued to be subcutaneously injected with CCl4 at the same time.The pathological changes of liver tissue were observed by HE and Masson staining.The levels of serum type Ⅳ collagen(Col Ⅳ),type Ⅲ procollagen(PC Ⅲ),hyaluronic acid(HA)and laminin(LN)were detected by ELISA.The protein expression levels of Wnt1,β-catenin and PPAR-γ in liver tissue were detected by Western Blot.The mRNA expression levels of Wnt1,β-catenin and PPAR-γ in liver tissue were detected by qPCR.Results Compared with the blank control group,the liver of the model group showed partial necrosis of liver cells,the normal hepatic lobule structure was destroyed,the arrangement of liver cell cords was disordered,the central vein or portal area was enlarged,and a large number of inflammatory cells infiltrated to form bridging necrosis.The collagen fibers in the central vein or portal area of the liver proliferated significantly,forming fibrous septa,and the fibrosis score were significantly increased(P＜0.05).The levels of serum Col IV,PC Ⅲ,HA and LN were significantly increased(P＜0.05).The protein and mRNA expression levels of Wnt1 and β-catenin in liver tissue were significantly increased(P＜0.05),and the protein and mRNA expression levels of PPAR-γ were significantly decreased(P＜0.05).Compared with the model group,the steatosis of hepatocytes in the positive control group and the medium-and high-dose groups of Chinese medicine was improved,and the necrosis and inflammatory cell infiltration were reduced.The degree of hepatitic fibrosis was improved,the liver collagen fibers were reduced,and the fibrosis score was significantly reduced(P＜0.05).The levels of serum Col Ⅳ,PC Ⅲ,HA and LN were significantly decreased(P＜0.05).The protein and mRNA expression levels of Wnt1 and β-catenin in liver tissue were significantly decreased(P＜0.05),and the protein and mRNA expression levels of PPAR-γ were significantly increased(P＜0.05).Conclusion Shuangzhu Kangxian Prescription has the effect of anti CCl4-induced hepatitic fibrosis in rats,and its mechanism may be related to the inhibition of Wnt/β-catenin signaling pathway and up-regulation of PPAR-γ expression.
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OBJECTIVE To investigate the effects of catalpol on H2O2-induced osteoblast injury and its mechanism. METHODS The osteoblasts MC3T3-E1 were separated into control group， model group， empty group （transfected with empty plasmid）， catalpol group （100 μmol/L）， catalpol+forkhead box O3 （FoxO3） overexpression group （100 μmol/L catalpol+ transfected with FoxO3 overexpression plasmid）. After catalpol treatment and transfection， except for control group， other groups were induced with H2O2 to establish osteoblast oxidative stress model. The cell viability， apoptotic rate， alkaline phosphatase （ALP） activity， optical density （OD） value of calcium nodule， mean fluorescence intensity （MFI） of reactive oxygen species （ROS）， antioxidant enzyme activity [superoxide dismutase （SOD）， catalase （CAT）]， the levels of inflammatory factors [interleukin-6 （IL-6）， IL-1β]， and the expressions of FoxO3/Wnt/β-catenin signaling pathway-related proteins were detected in each group. RESULTS Compared with the control group， the cell viability， ALP activity， OD value of calcium nodule， activities of antioxidant enzyme， and the protein expressions of Wnt and β-catenin were decreased significantly in the model group， while apoptotic rate， MFI levels of ROS， inflammatory factor levels and the protein expression of FoxO3 were all increased significantly （P＜0.05）. Compared with the model group， above indicators of the empty group had no significant change （P＞0.05）， while those of catalpol group were reversed significantly （P＜0.05）. Compared with the catalpol group， the reversal effect of the changes in the above indicators was significantly weakened in the catalpol+FoxO3 overexpression group cells （P＜0.05）. CONCLUSIONS Catalpol can activate Wnt/β-catenin signaling pathway by down-regulating FoxO3， thereby inhibiting H2O2-induced MC3T3-E1 oxidative stress and inflammation reaction， enhancing cell viability and osteogenic differentiation activity， and alleviating apoptosis injury.
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ObjectiveTo observe the effects of the kidney-tonifying and blood-activating prescription on the Wnt/β-catenin signaling pathway and uterine spiral artery remodeling in a mouse model of recurrent miscarriage and to explore its underlying mechanism. MethodA mouse model of normal pregnancy was established by mating CBA/J mice with BALB/c mice. A mouse model of recurrent miscarriage was established by mating CBA/J mice with DBA/2 mice. The modeled mice of recurrent miscarriage were randomized into model， dydrogesterone， and low- and high-dose Chinese medicine groups. The mice in normal pregnancy were used as the control group. Each group consisted of 10 mice， and the drug administration lasted for 14 days. After the treatment， the embryo absorption rate of each group was recorded. Hematoxylin-eosin （HE） staining was employed to observe the pathological morphology of the uterine decidua， and the physiological transformation rate of spiral arteries （SPA） was evaluated. Real-time polymerase chain reaction （Real-time PCR） and Western blot were performed to determine the mRNA and protein levels， respectively， of matrix metalloproteinases （MMP）-2， MMP-9， vascular endothelial growth factor （VEGF）， and Wnt/β-catenin signaling pathway. ResultCompared with the control group， the model group presented increased embryo absorption rate （P<0.05）， decreased physiological transformation rate of uterine SPA （P<0.05）， cellular swelling， degeneration， and disordered arrangement in the uterine decidua tissue， and down-regulated mRNA and protein levels of key factors involved in SPA remodeling （MMP-2， MMP-9， VEGF） and the Wnt/β-catenin signaling pathway （Wnt2， β-catenin， Cyclin D1， c-Myc） （P<0.05）. Compared with the model group， both the low- and high-dose Chinese medicine reduced embryo absorption rate （P<0.05）， increased SPA physiological transformation rate （P<0.05）， improved uterine decidua tissue morphology， and increased decidua vessel count. Furthermore， they up-regulated the mRNA and protein levels of MMP-2， MMP-9， VEGF， and proteins in the Wnt/β-catenin signaling pathway （P<0.05）. ConclusionRecurrent miscarriage is associated with impaired uterine spiral artery remodeling. The kidney-tonifying and blood-activating prescription can promote uterine spiral artery remodeling by activating the Wnt/β-catenin signaling pathway and promoting the expression of VEGF， MMP-2， and MMP-9， thus treating recurrent miscarriage.
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Lung cancer is one of the most common malignant tumors in the world， with its morbidity and mortality ranking at the top. The early symptoms are not obvious， and the biological structure is complex， so many patients have missed the optimal treatment time. At present， the treatment of lung cancer in modern medicine is dominated by first-line chemotherapy and surgical treatment with platinum-containing regimen， which has relatively large side effects， poor prognosis， and a high risk of metastasis and recurrence. With the gradual rise of targeted therapy and immunotherapy for lung cancer， the overall recovery of patients with lung cancer is still poor and the survival rate is low， despite more abundant treatment methods. From the perspective of holistic concept and syndrome differentiation， traditional Chinese medicine （TCM） plays an important role in the prognosis of tumor patients， with many targets， a wide range and light toxic and side effect. Modern studies have shown that the occurrence and development of lung cancer are closely related to the abnormality of multiple signaling pathways， and the Wnt/β-catenin signaling pathway， as one of the most important pathways in cancer， is involved in the whole process of lung cancer development by regulating the expression of related signaling proteins and genes. In recent years， many studies have confirmed that TCM monomers and TCM compounds can inhibit the epithelial-mesenchymal transition （EMT） process of lung cancer and the activity of lung cancer stem cells （LCSCs） by regulating the Wnt/β-catenin signaling pathway， induce lung cancer cell apoptosis， inhibit the proliferation， invasion and migration of lung cancer cells， and thus play an anti-lung cancer role. In recent years， research in this field has made breakthroughs， but there is a lack of systematic reviews and summaries. Thus， this paper reviewed relevant literature worldwide to analyze and interpret the mechanism of TCM intervention in the Wnt/β-catenin signaling pathway against lung cancer. The TCM monomers targeted to regulate this signaling pathway were summarized in four categories： promoting blood circulation for removing blood stasis， clearing heat and removing dampness， clearing heat and removing toxicity， and awakening the spirit. TCM compounds included Buzhong Yiqitang， Xuefu Zhuyutang， et al. This study aims to provide new ideas for clinical research and drug development for lung cancer.
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R-spondin2 (Rspo2) is a member of protein family RSPOs, which can be coupled to receptor 4/5 (leucine-rich repeat-containing g protein-coupled receptor 4/5, LGR4/5), cell surface transmembrane E3 ubiquitin ligase ZNRF3/RNF43 (zinc and ring finger 3/ring finger protein 43), heparan sulfate proteoglycan (heparan sulfate proteoglycans, HSPGs) and the IQ motif (IQ gap 1) containing GTP enzyme activating protein 1, regulating the Wnt/β-catenin signaling pathway, which is the most widely studied signaling pathway and directly related to basic bone biology. Any problem in this pathway may have an impact on bone regulation. In recent years, it has been found that Rspo2 can act on osteoblast, osteoclast and chondrocytes through Wnt/β-catenin, and take part in occureace and development of some bone diseases such as ossification of the posterior longitudinal ligament (OPLL), osteoarthritis (OA) and rheumatoid arthritis (RA), so the study of Rspo2 may become a new therapeutic direction for bone-related diseases. Based on the latest research progress, this paper reviews the structure and main functions of Rspo2, the mechanism of Rspo2 regulating Wnt/β-catenin signaling pathway and its influence on skeletal system, in order to provide new ideas and ways for the prevention and treatment of bone-related diseases.
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Objective To investigate the effects of calcium and integrin-binding protein 1 (CIB1) on the cell proliferation, invasion, apoptosis, and migration of triple-negative breast cancer cells and its possible mechanism. Methods MDA-MB-231 and MDA-MB-468 cells were divided into CIB1-knockdown(infected with CRISPR/Cas9 lentivirus) and negative-control groups. Cell Counting Kit-8 (CCK-8) and 5-ethynyl-2′-deoxyuridine (EdU) assays were performed to detect cell proliferation. Cell apoptosis was determined through flow cytometry. Scratch and Transwell experiments were conducted to measure the migration and invasion abilities of cells. The mRNA and protein expression levels of β-catenin, adenomatous polyposis coli (APC), glycogen synthase kinase 3β (GSK-3β), and c-myc were detected via real-time quantitative polymerase chain reaction and Western blot. Results Compared with the negative-control group, the CIB1-knockdown group showed decreased cell proliferation, invasion, and migration (P<0.05) and increased cell apoptosis (P<0.05). The mRNA and protein expressions of β-catenin, APC, and c-myc decreased (P<0.05), and that of GSK-3β increased (P<0.05). Conclusion CIB1 knockdown can inhibit cell proliferation, invasion, and migration and promote the apoptosis of breast cancer cells. Its mechanism may be related to the inhibition of Wnt/β-catenin signaling pathway.
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Diabetic kidney disease （DKD） is a common complication of diabetes and a leading cause of end-stage kidney disease. Its pathogenesis is complex， and it presents a significant challenge in treatment， gradually becoming a major global public health issue. One of the main pathological changes in DKD is tubulointerstitial fibrosis， clinically characterized by proteinuria and declining kidney function， which severely impacts the daily life of patients. Currently， western medicine commonly uses methods such as controlling blood sugar and blood pressure， and reducing proteinuria to treat DKD， but the efficacy is unsatisfactory， and there are many side effects. As reported， traditional Chinese medicine （TCM） treatment for DKD has many advantages， such as low cost， significant efficacy， and minimal adverse reactions. More researchers focusing on DKD are turning their attention to TCM， and progress has been made in related studies both in China and abroad. The Wnt/β-catenin signaling pathway is relatively evolutionarily conserved and plays a crucial role in normal biological development and the entire life process. Studies have demonstrated that abnormal activation of the Wnt/β-catenin signaling pathway is related to renal fibrosis， which coincides with TCM theory of "collateral diseases". By reviewing relevant literature， this article reviewed the Wnt/β-catenin signaling pathway and its role in DKD and summarized the research status of TCM monomers， single drug extracts， and TCM formulas in improving renal fibrosis and treating DKD through the improvement of glomerular mesangial cells， renal tubular epithelial cells， and podocyte injury， aiming to provide new ideas and directions for TCM treatment of DKD.
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OBJECTIVE To explore the intervention effect and related mechanism of Tongxinluo capsule on renal fibrosis in rats with diabetic nephropathy （DN）. METHODS Eight rats were selected as control group （ordinary feed）， the remaining rats were given high-glucose and high-fat diet combined with ip injection of streptozotocin （35 mg/kg） to induce DN model. Model rats were randomly divided into model group （purified water）， irbesartan group （positive control， 14.12 mg/kg） and Tongxinluo capsule group （0.3 g/kg）， including 12 rats in the model group and 11 rats for each of the other two groups. All groups were given relevant medicine or water intragastrically， once a day， for 16 consecutive weeks. After the last medication， fasting blood glucose and 24 h urinary total protein （24 h UTP） were detected. Pathological changes in renal cortex of rats in each group were observed. Serum levels of tissue-type plasminogen activator （PA） and plasminogen activator inhibitor 1 （PAI-1） were measured. mRNA expressions of transforming growth factor-β（1 TGF-β1）， type Ⅳ collagen（COL-Ⅳ）， Wnt4 and β-catenin in renal cortex of rats were detected. The protein depositions or expressions of TGF-β1， COL-Ⅳ， focal adhesion kinase （FAK）， integrin-linked kinase （ILK）， E-cadherin， PA， PAI-1， Wnt4 and β-catenin in renal cortex of rats were observed or determined. RESULTS Compared with model group， 24 h UTP of rats in Tongxinluo capsule group were all significantly reduced （P＜0.05）； pathological damage and fibrosis of renal cortex were relieved； the expression of PA in serum and renal cortex was significantly increased， while PAI-1 level was significantly reduced （P＜0.05）； the depositions of COL-Ⅳ and TGF-β1 in renal cortex were all reduced， and corresponding mRNA expression was decreased significantly （P＜0.05）； the depositions of ILK and FAK were decreased， while the deposition of E-cadherin was increased； protein and mRNA expressions of Wnt4 and β-catenin were significantly reduced （P＜0.05）. CONCLUSIONS Tongxinluo capsule can relieve pathological damage to renal tissue and renal fibrosis of DN model rats， and reduce extracellular matrix deposition. The mechanism may be related to regulation of fibrinolytic system activity， the decrease of ILK and FAK expression， and inhibition of Wnt/β-catenin signaling pathway.
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OBJECTIVE@#To observe the effect of wheat-grain moxibustion at "Dazhui" (GV 14), "Zusanli" (ST 36) and "Sanyinjiao" (SP 6) on Wnt/β-catenin signaling pathway in bone marrow cell in mice with bone marrow inhibition, and to explore the possible mechanism of wheat-grain moxibustion in treating bone marrow inhibition.@*METHODS@#Forty-five SPF male CD1(ICR) mice were randomly divided into a blank group, a model group and a wheat-grain moxibustion group, 15 mice in each group. The bone marrow inhibition model was established by intraperitoneal injection of 80 mg/kg of cyclophosphamide (CTX). The mice in the wheat-grain moxibustion group were treated with wheat-grain moxibustion at "Dazhui" (GV 14), "Zusanli" (ST 36) and "Sanyinjiao" (SP 6), 3 moxa cones per acupoint, 30 s per moxa cone, once a day, for 7 consecutive days. The white blood cell count (WBC) was measured before modeling, before intervention and 3, 5 d and 7 d into intervention. After intervention, the general situation of mice was observed; the number of nucleated cells in bone marrow was detected; the serum levels of interleukin-3 (IL-3), interleukin-6 (IL-6) and granulocyte macrophage colony stimulating factor (GM-CSF) were measured by ELISA; the protein and mRNA expression of β-catenin, cyclinD1 and C-Myc in bone marrow cells was measured by Western blot and real-time PCR method.@*RESULTS@#Compared with the blank group, the mice in the model group showed sluggish reaction, unstable gait, decreased body weight, and the WBC, number of nucleated cells in bone marrow as well as serum levels of IL-3, IL-6, GM-CSF were decreased (P<0.01), and the protein and mRNA expression of β-catenin, cyclinD1 and C-Myc was decreased (P<0.01). Compared with the model group, the mice in the wheat-grain moxibustion group showed better general condition, and WBC, the number of nucleated cells in bone marrow as well as serum levels of IL-3, IL-6, GM-CSF were increased (P<0.01, P<0.05), and the protein and mRNA expression of β-catenin, cyclinD1 and C-Myc was increased (P<0.05).@*CONCLUSION@#Wheat-grain moxibustion shows therapeutic effect on bone marrow inhibition, and its mechanism may be related to activating Wnt/β-catenin signaling pathway in bone marrow cells, improving bone medullary hematopoiesis microenvironment and promoting bone marrow cell proliferation.
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Animales , Masculino , Ratones , beta Catenina/metabolismo , Médula Ósea/fisiopatología , Células de la Médula Ósea/fisiología , Factor Estimulante de Colonias de Granulocitos y Macrófagos/metabolismo , Interleucina-3/metabolismo , Interleucina-6/metabolismo , Ratones Endogámicos ICR , Moxibustión/métodos , ARN Mensajero/metabolismo , Triticum , Vía de Señalización Wnt , HematopoyesisRESUMEN
OBJECTIVE@#To investigate the mechanism by which fibroblasts with high WNT2b expression causes intestinal mucosa barrier disruption and promote the progression of inflammatory bowel disease (IBD).@*METHODS@#Caco-2 cells were treated with 20% fibroblast conditioned medium or co-cultured with fibroblasts highly expressing WNT2b, with the cells without treatment with the conditioned medium and cells co-cultured with wild-type fibroblasts as the control groups. The changes in barrier permeability of Caco-2 cells were assessed by measuring transmembrane resistance and Lucifer Yellow permeability. In Caco-2 cells co-cultured with WNT2b-overexpressing or control intestinal fibroblasts, nuclear entry of β-catenin was detected with immunofluorescence assay, and the expressions of tight junction proteins ZO-1 and E-cadherin were detected with Western blotting. In a C57 mouse model of dextran sulfate sodium (DSS)-induced IBD-like enteritis, the therapeutic effect of intraperitoneal injection of salinomycin (5 mg/kg, an inhibitor of WNT/β-catenin signaling pathway) was evaluated by observing the changes in intestinal inflammation and detecting the expressions of tight junction proteins.@*RESULTS@#In the coculture system, WNT2b overexpression in the fibroblasts significantly promoted nuclear entry of β-catenin (P < 0.01) and decreased the expressions of tight junction proteins in Caco-2 cells; knockdown of FZD4 expression in Caco-2 cells obviously reversed this effect. In DSS-treated mice, salinomycin treatment significantly reduced intestinal inflammation and increased the expressions of tight junction proteins in the intestinal mucosa.@*CONCLUSION@#Intestinal fibroblasts overexpressing WNT2b causes impairment of intestinal mucosal barrier function and can be a potential target for treatment of IBD.
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Humanos , Ratones , Animales , Células CACO-2 , beta Catenina/metabolismo , Medios de Cultivo Condicionados/farmacología , Uniones Estrechas/metabolismo , Mucosa Intestinal , Enfermedades Inflamatorias del Intestino , Proteínas de Uniones Estrechas/metabolismo , Inflamación/metabolismo , Fibroblastos/metabolismo , Ratones Endogámicos C57BL , Glicoproteínas/metabolismo , Proteínas Wnt/farmacología , Receptores Frizzled/metabolismoRESUMEN
AIM: To scrutinize the role of the Wnt/β-catenin signaling pathway in the epithelial-mesenchymal transition(EMT)of lens epithelial cells under hypoxic conditions, and to further analyze the effect of Dickkopf-1(DKK-1)expression on EMT of lens epithelial cells.METHODS: Human lens epithelial cells(HLEB3 cells)were propagated in vitro and then separated into two groups: one exposed to standard oxygen levels, added DMEM culture solution containing 10% FBS(normoxic group)and another subjected to low oxygen levels(hypoxic group). The hypoxic condition was emulated by applying a concentration of 100 μmol/L cobalt chloride(CoCl2)for 6, 12, 24, and 48h. The utilization of immunofluorescence staining enabled the detection of Wnt3a and DKK-1 expressions, along with the expression and localization of β-catenin protein in these groups. The expression of DKK-1 mRNA was discerned by quantitative real-time polymerase chain reaction(qRT-PCR).RESULTS: Immunofluorescence assays indicated an escalating trend in the Wnt3a and DKK-1 protein expression, which corresponded with the increasing duration of hypoxia. Likewise, an intensified nuclear accumulation of β-catenin protein was observed to be directly proportional to the length of hypoxia treatment. The qRT-PCR demonstrated that the difference in DKK-1 mRNA expression between the normoxic group and the group exposed to hypoxia for 6h was not statistically significant(P>0.05), whereas the DKK-1 mRNA expression of the 12, 24, and 48h hypoxia groups were significantly increased(P<0.001).CONCLUSION: Hypoxia can activate Wnt/β-catenin pathway in lens epithelial cells and induce the expression of DKK-1, thus regulating the Wnt/β-catenin pathway and affecting the EMT process.
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Steroid-induced necrosis of the femoral head （SNOFH） is a common orthopedic disease，which is difficult to cure and has poor clinical prognosis. The number of SNOFH patients in China is still increasing year by year，which seriously threatens human health. Long-term non-standard or short-term extensive use of hormone （GC） is an important reason for the occurrence of this disease. At present，SNOFH is mostly treated by surgical methods such as hip replacement，which has limitations of great harm to patients and high cost. In recent years，with the continuous deepening and innovation of traditional Chinese medicine（TCM） research，the use of TCM to treat SNOFH has been widely used in clinical practice. The main TCM pathogenesis of SNOFH is kidney deficiency and blood stasis. Therefore，TCM monomer and compound compound of tonifying kidney and promoting blood circulation are used to treat SNOFH. And TCM has obvious therapeutic effect，small side effects，less cost and other advantages. Glycoprotein/beta chain protein secretion （Wnt/beta- catenin） signaling pathway as a classic signaling pathway is closely related to the bone，between its by promoting bone marrow mesenchymal stem cell update，enhance the activity of osteoblast and suppress the apoptosis，which adjust the metabolic balance of bone tissue，increase bone density，will play an important role in the process of bone formation. In recent years，the use of TCM monomers and compounds to regulate Wnt/β-catenin signaling pathway to accelerate bone marrow mesenchymal stem cells，promote their transformation into osteoblasts，and maintain bone metabolic balance mechanism to treat SNOFH has become a new research hotspot. This article reviews the research progress of TCM in the prevention and treatment of SNOFH by regulating Wnt/β-catenin signaling pathway，in order to provide reference for the application of TCM in the treatment of SNOFH.
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OBJECTIVE To investigate whether icari-in(ICA)plays a neuroprotective role by improving glyco-lytic function through activating Wnt/β-catenin signaling pathway.METHODS HT22 cells were treated with Aβ25-35 for 24 h to establish AD cell model,ICA was added in 2 h before Aβ25-35 and the DKK1(a specific inhibitor of the Wnt signaling pathway)was added in 0.5 h before ICA.Pharmacodynamic study:HT22 cells were divided into control group,ICA group(ICA 10 μmol·L-1),model group(Aβ25-3520 μmol·L-1),model + ICA group(Aβ25-3520 μmol·L-1 +ICA 2.5,5,10 μmol·L-1);Mechanism study:HT22 cells were divided into control group,model group,Aβ25-35+ICA 10 μmol·L-1 group,Aβ25-35+DKK1 group,Aβ25-35+DKK1+ ICA group.The cell viability was detected by MTT assay and the cell morphology was obtained by microscope,the lactate content was detected by lactate assay,the ATP content was measured with the chemiluminescence method,the expression levels of HK1,PKM1 and the pro-tein expression of molecules related to the Wnt/β-catenin signaling pathway(Wnt3a,GSK3β,pGSK3β Try216,pGSK3β Ser9,β-catenin,pβ-catenin Ser33/37 Thr41,Active β-catenin and nuclear β-catenin)was assayed by Western blotting.The nuclear translocation of β-catenin was observed by immunofluorescent staining.RESULTS Compared with the control group,the viability of cells in the model group was reduced,the morphology of cells was significantly damaged,the ATP content and lactate content were significantly decreased,and the glycolytic key enzymes:the protein levels of HK1,PKM1 and the protein levels of Wnt3a,pGSK3β Ser9,active β-catenin and nuclear β-catenin were significantly reduced,and the phosphorylation levels of β-catenin Ser33/37 Thr41 were significantly increased.Compared with the model group,the cell morphology was significantly improved and the viability was significantly increased,the ATP and lactate content were significantly increased,the expressions of HK1,PKM1 and Wnt3a,pGSK3β Ser9,active β-catenin and nuclear β-catenin protein were significantly upregulat-ed,and the phosphorylation levels of β-catenin Ser33/37 Thr41 were significantly reduced after ICA treatment.However,when the canonical Wnt signaling was inhibited by DKK1,the above effects of ICA on glycolysis were abolished.CONCLUSION ICA exerts neuroprotective effects on Aβ25-35-induced HT22 cell injury by enhancing the glycolysis function through the activation of the Wnt/β-catenin signaling pathway.
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Taste is a critical sensory function for human as it supports sustenance and alerts the body to toxins. Taste dysfunction is a common side effect of radiotherapy for the head and neck cancers, which is often accompanied by oral mucositis in the early stage. It is associated with anorexia, anxiety and depression, leading to declined quality of life and treatment tolerance. The incidence of radiation-induced taste dysfunction is high, and its clinical manifestations include increased taste threshold, tastelessness, and persistent bitter, sour or metallic taste, which exert significant effect upon the quality of life. At present, effective therapeutic measures for radiation-induced taste dysfunction are still lacking. In this article, research progresses on clinical characteristics and the potential mechanisms of radiation-induced taste dysfunction were reviewed, aiming to provide reference for the mechanism, prevention and treatment for taste dysfunction.
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Objective:To investigate the role of miR-765 in papillary thyroid carcinoma (PTC) cells and further uncover the associated signaling mechanism.Methods:qPCR was used to assess miR-765 expression in normal human thyroid cell line (Nthy-ori 3-1) and human PTC cell lines (B-CPAP and TPC-1). PTC cells were divided into blank control group (BC) without special treatment, negative control group (NC) that was transfected with negative control sequences, and miR-mimic group that was transfected with miR-mimic. Transfection with miR-mimic was used to up-regulate the expression of miR-765 in PTC cells. CCK-8, plate colony formation, wound-healing, and Transwell invasion assays were used to assess the proliferation, migration, and invasion of PTC cells, respectively. Western blot assay was used to assess the level of nuclear β-catenin, the key protein of the Wnt/β-catenin pathway, in PTC cells.Results:The level of miR-765 expression of PTC cells was significantly lower than that of Nthy-ori 3-1 cells (B-CPAP, P=0.0003; TPC-1, P=0.0003). Transfection with miR-mimic significantly up-regulated miR-765 expression in PTC cells (B-CPAP, P<0.0001; TPC-1, P<0.0001). Results of CCK-8 assay (B-CPAP, P<0.05; TPC-1, P<0.05), plate colony formation assay (B-CPAP, P=0.0001; TPC-1, P<0.0001), wound-healing assay, and Transwell invasion assay (B-CPAP, P=0.001; TPC-1, P=0.0014) showed that up-regulating the expression of miR-765 significantly inhibited the proliferation, migration, and invasion of PTC cells. Western blot results showed that up-regulating the expression of miR-765 significantly reduced nuclear β-catenin (B-CPAP, P=0.0039; TPC-1, P=0.0004) . Conclusion:up-regulating the expression of miR-765 inhibits the proliferation, migration, and invasion of PTC cells and the Wnt/β-catenin signaling pathway, which not only proposes miR-765 as a novel potential therapeutic target for PTC, but also further revealed the associated mechanism.
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Objective To investigate the mechanism of metformin on oxygen-glucose deprivation/reoxygenation(OGD/R)injury in U251 cells.Methods Human glioma cell line U251 cells were cultured and divided into 6groups:blank control group,model group,metformin medium dose group,metformin high dose group,agonist group(Wnt3a,Wnt/β-catenin signaling pathway agonist),inhibitor group(metformin+XAV939,Wnt/β-catenin signaling pathway inhibitor).Except for the blank control group,the cells in the other groups were subjected to OGD/R for 2h and then reperfusion for 24h to establish the OGD/R model.The animals were treated with met-formin,Wnt3a and XAV939 24h before modeling.Cell viability and toxicity were detected by CCK-8method and lactate dehydrogenase(LDH)assay.ROS formation was detected by DHE staining.Glutathione peroxidase(GSH-Px),superoxide dismutase(SOD)malond-ialdehyde(MDA),interleukin-6(IL-6),inducible nitric oxide synthase(iNOS)and tumor necrosis factor-α(TNF-α)were de-tected by enzyme-linked immunoadsordent assay(ELISA).The protein expression levels of β-catenin,cyclin D1,p-GSK-3β(Ser9)and GSK-3β were detected by Western blot.Results Compared with blank control group,LDH,ROS,MDA,IL-6,iNOS and TNF-α in model group,metformin group,agonist group and inhibitor group were significantly increase.The relative expression lev-els of SOD,GSH-Px,β-catenin,cyclin D1,p-GSK-3β(Ser9)and cell viability were significantly decreased.Compared with mod-el group,the levels of LDH,ROS,MDA,IL-6,iNOS and TNF-α in metformin group and agonist group were significantly decreased,while the relative expression levels of SOD,GSH-Px,β-catenin,cyclin D1,p-GSK-3β(Ser9)and the cell viability were signifi-cantly increased.Compared with metformin group,LDH,ROS,MDA,IL-6,iNOS and TNF-α in metformin group and inhibitor group were significantly increase,the relative expression levels of SOD,GSH-Px,β-catenin,cyclin D1,p-GSK-3 β(Ser9)and the cell viability were significantly decreased.Conclusion Metformin may play a protective role in OGD/R of U251 cells through Wnt/β-cate-nin signaling pathway.