scholarly journals The Role of Neutrophil Elastase in Aortic Valve Calcification

2022 ◽  
Author(s):  
Yan Liu ◽  
Peng Jiang ◽  
Liqin An ◽  
Mengying Zhu ◽  
Jin Li ◽  
...  
Keyword(s):  
Aortic Valve ◽  
Western Blot ◽  
Osteogenic Differentiation ◽  
Neutrophil Elastase ◽  
Calcium Deposition ◽  
Enzyme Linked Immunosorbent Assay ◽  
Osteogenic Medium ◽  
Rt Pcr ◽  
Alizarin Red ◽  
Valve Calcification

Abstract Background: Calcific aortic valve disease (CAVD) is the most commonly valvular disease in the western countries initiated by inflammation and abnormal calcium deposition. Currently, there is no clinical drugs for CAVD. Neutrophil elastase(NE) plays a causal role in inflammation and participates actively in cardiovascular diseases. However, the effects of NE on valve calcification remains unclear. So we next explore whether it is involved in valve calcification and the molecular mechanisms involved.Methods: NE expression and activity in calcific aortic valve stenosis (CAVS) patients (n=58) and healthy patients (n=30) were measured by enzyme-linked immunosorbent assay (ELISA), western blot and immunohistochemistry (IHC). Porcine aortic valve interstitial cells (pVICs) were isolated and used in vitro expriments. The effects of NE on pVICs inflammation, apoptosis and calcification were detected by hochest 33258 staining, MTT assay, reverse transcription polymerase chain reaction (RT-PCR) and western blot. The effects of NE knockdown and NE activity inhibitor Alvelestat on pVICs inflammation, apoptosis and calcification under osteogenic medium induction were also detected by RT-PCR, western blot, alkaline phosphatase staining and alizarin red staining. Changes of Intracellular signaling pathways after NE treatment were measured by western blot.Results: The level and activity of NE were evaluated in patients with CAVS and calcified valve tissues. NE promoted inflammation, apoptosis and phenotype transition in pVICs in the presence or absence of osteogenic medium. Under osteogenic medium induction, NE silencing or NE inhibitor Alvelestat both suppressed the osteogenic differentiation of pVICs. Mechanically, NE played its role in promoting osteogenic differentiation of pVICs by activating the NF-κB and AKT signaling pathway.Conclusions: Collectively, NE is highly involved in the pathogenesis of valve calcification. Targeting NE such as Alvelestat may be a potential treatment for CAVD.

scholarly journals High Glucose Enhances the Odonto/Osteogenic Differentiation of Stem Cells from Apical Papilla via NF-KappaB Signaling Pathway

2019 ◽  
Vol 2019 ◽  
pp. 1-10 ◽  
Author(s):  
Yanping Wang ◽  
Yanqiu Wang ◽  
Yadie Lu ◽  
Jinhua Yu
Keyword(s):  
Stem Cells ◽  
Western Blot ◽  
Osteogenic Differentiation ◽  
Real Time ◽  
Signaling Pathway ◽  
Mtt Assay ◽  
High Glucose ◽  
Rt Pcr ◽  
Alizarin Red ◽  
Apical Papilla

Objective. The transport and metabolism of glucose are important during mammalian development. High glucose can mediate the biological characteristics of mesenchymal stem cells (MSCs). However, the role of high glucose in the odonto/osteogenic differentiation of stem cells from apical papilla (SCAPs) is unclear. Materials and Methods. SCAPs were isolated and identified in vitro. Then, SCAPs were cultured in normal α-MEM and high glucose α-MEM separately. MTT assay was applied to observe the proliferation of SCAPs. ALP activity, alizarin red staining, real-time RT-PCR, and western blot were used to detect the odonto/osteogenic capacity of SCAPs as well as the participation of NF-κB pathway. Results. SCAPs in 25mmol/L glucose group expressed the maximum proteins of RUNX2 and ALP as compared with those in 5, 10, and 15 mmol/L groups. MTT assay showed that 25 mmol/L glucose suppressed the proliferation of SCAPs. ALP assay, alizarin red staining, real-time RT-PCR, and western blot showed 25 mmol/L high glucose can obviously enhance the odonto/osteogenic capacity of SCAPs. Moreover, the NF-κB pathway was activated in 25mmol/L glucose-treated SCAPs and the odonto/osteogenic differentiation was inhibited following the inhibition of NF-κB signaling pathway. Conclusions. High glucose can enhance the odonto/osteogenic capacity of SCAPs via NF-κB pathway.

scholarly journals Baicalein inhibits inflammatory response and promotes osteogenic activity in periodontal ligament cells challenged with lipopolysaccharides

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Manman Ren ◽  
Ya Zhao ◽  
Zhiqi He ◽  
Jian Lin ◽  
Chuchu Xu ◽  
...  
Keyword(s):  
Inflammatory Response ◽  
Western Blot ◽  
Osteogenic Differentiation ◽  
Periodontal Ligament ◽  
Mapk Signaling ◽  
Periodontal Ligament Cells ◽  
Enzyme Linked Immunosorbent Assay ◽  
Alizarin Red ◽  
Osteogenic Markers ◽  
Anti Inflammatory

Abstract Background Periodontitis is a chronic infection initiated by oral bacterial and their virulence factors, yet the severity of periodontitis is largely determined by the dysregulated host immuno-inflammatory response. Baicalein is a flavonoid extracted from Scutellaria baicalensis with promising anti-inflammatory properties. This study aims to clarify the anti-inflammatory and osteogenic effects of baicalein in periodontal ligament cells (PDLCs) treated with lipopolysaccharides (LPS). Methods Human PDLCs were incubated with baicalein (0–100 μM) for 2 h prior to LPS challenge for 24 h. MTT analysis was adopted to assess the cytoxicity of baicalein. The mRNA and protein expression of inflammatory and osteogenic markers were measured by real-time polymerase chain reaction (PCR), western blot and enzyme-linked immunosorbent assay (ELISA) as appropriate. Alkaline phosphatase (ALP) and Alizarin red S (ARS) staining were performed to evaluate the osteogenic differentiation of PDLCs. The expression of Wnt/β-catenin and mitogen-activated protein kinase (MAPK) signaling related proteins was assessed by western blot. Results MTT results showed that baicalein up to 100 μM had no cytotoxicity on PDLCs. Baicalein significantly attenuated the inflammatory factors induced by LPS, including interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), matrix metalloprotein-1 (MMP-1), MMP-2 and monocyte chemoattractant protein 1 (MCP-1) at both mRNA and protein level. Moreover, MAPK signaling (ERK, JNK and p38) was significantly inhibited by baicalein, which may account for the mitigated inflammatory response. Next, we found that baicalein effectively restored the osteogenic differentiation of LPS-treated PDLCs, as shown by the increased ALP and ARS staining. Accordingly, the protein and gene expression of osteogenic markers, namely runt-related transcription factor 2 (RUNX2), collagen-I, and osterix were markedly upregulated. Importantly, baicalein could function as the Wnt/β-catenin signaling activator, which may lead to the increased osteoblastic differentiation of PDLCs. Conclusions With the limitation of the study, we provide in vitro evidence that baicalein ameliorates inflammatory response and restores osteogenesis in PDLCs challenged with LPS, indicating its potential use as the host response modulator for the management of periodontitis.

P5991Cellular mechanisms of aortic valve calcification in primary human valvular interstitial cell culture

2019 ◽  
Vol 40 (Supplement_1) ◽  
Author(s):  
E Zhiduleva ◽  
O Irtyuga ◽  
P Murtazalieva ◽  
J Sibagatullina ◽  
A Shishkova ◽  
...  
Keyword(s):  
Aortic Valve ◽  
Aortic Stenosis ◽  
Osteogenic Differentiation ◽  
Bone Morphogenetic Protein 2 ◽  
Osteogenic Potential ◽  
Orthotopic Heart Transplantation ◽  
Control Group ◽  
Alizarin Red ◽  
Valve Calcification ◽  
Tricuspid Aortic Valve

Abstract Background Aortic stenosis (AS) is the most common acquired valvular heart disease. Calcification of the aortic valve (AV) cusps is the main pathogenetic mechanism of AS formation, however triggering and progression mechanisms of it are not fully understood. Aortic valve interstitial cells (AVIC) are one of the main cell populations responsible for the AV structure and homeostasis. The aim of the study was to characterize the ability of AVIC to osteogenic differentiation using the model of the primary culture of AVIC of patients with aortic stenosis and assess the expression of genes involved in osteogenesis (RUNX2, BMPG2, SPRY1, SOX9, CTNNB1, POSTN, OPG and OPN). Materials and methods The study was carried out on primary human AVIC culture obtained by enzymatic dissociation of experimental valve specimens from patients with AS (n=15) and the control specimens from the orthotopic heart transplantation recipients (n=10). To assess the osteogenic potential of AVIC, the routine stem cell osteodifferentiation protocol based on the culture medium with addition of inductors (10 mM β-glycerophosphate, 0.1 μM dexamethasone, and 50 μg/ml ascorbic acid) was used. Calcium deposits were demonstrated by Alizarin Red staining. Analysis of the expression of osteogenic differentiation genes, such as RUNX2 (runtrelated transcription factor 2), BMP2 (bone morphogenetic protein 2), SPRY1 (Sprouty RTK signaling antagonist 1), SOX-9 (SRY-box9), CTNNB1 (β-catenin1), POSTN (periostin), OPG (osteoprotegerin), and OPN (osteopontin), was performed by real-time PCR. Results The inductors of osteogenic differentiation provoked greater mineralization of AVIC cultures derived from the patients with AS than that observed in control group (p=0.0003). The expression of RUNX2 and SPRY1 in non-differentiated cells was reduced compared with control cells in patients with bicuspid AV (p=0.02). After 21 days of the osteogenic induction, the expression of RUNX2 and SPRY1 increased in all three groups. At the same time, the expression of SPRY1 was lower in the group with bicuspid AV compared with tricuspid AV (p=0.007). The expression level of BMP2 did not differ between groups in unstimulated AVIC, however, it increased after osteogenic differentiation in the group of patients with tricuspid AV (p=0.017). OPN expression was higher in cells from tricuspid AV, while OPG expression was reduced in patients with both bicuspid and tricuspid AV (p<0.01), No differences were found between the groups) for the remaining genes (POSTIN, CTNNB1, SOX9) before and after stimulation with an osteogenic medium. Conclusions The osteogenic potential of AVIC is increased in patients with aortic stenosis. The gene expression profile of osteogenic differentiation differs in patients with a bicuspid and tricuspid aortic valve. Damage of protective mechanisms may be a potential mechanism for accelerated valve calcification mostly in patients with tricuspid aortic valve. Acknowledgement/Funding Russian Foundation for Basic Research, project 18-015-00016

scholarly journals Curculigoside promotes osteogenic differentiation of ADSCs to prevent ovariectomized-induced osteoporosis

2021 ◽  
Vol 16 (1) ◽  
Author(s):  
Wei-Li You ◽  
Zheng-Long Xu
Keyword(s):  
Bone Loss ◽  
Western Blot ◽  
Osteogenic Differentiation ◽  
Signaling Pathway ◽  
Akt Signaling ◽  
Calcium Deposition ◽  
Phenolic Glycoside ◽  
Akt Signaling Pathway ◽  
Alizarin Red ◽  
Alp Activity

Abstract Background Curculigoside is a natural phenolic glycoside compound produced by Curculigo orchioides Gaertn. This study aimed to explore the effects of curculigoside in promoting the osteogenic differentiation of adipose-derived stem cells (ADSCs) as well as the underlying mechanism. Methods ADSCs were treated with curculigoside at different concentrations (0 μmol/L, 1 μmol/L, 2.5 μmol/L, 5 μmol/L, 10 μmol/L, and 20 μmol/L), and cell viability was assessed by CCK-8 assay. Then, the alkaline phosphatase (ALP) activity was determined, and alizarin red S (ARS) staining was performed to measure the extracellular mineralization of curculigoside. Information about protein-chemical interactions is provided by the search tool for interactions of chemicals (STITCH) database. Then, LY294002 was administered to explore the mechanism by which curculigoside promotes the osteogenic differentiation of ADSCs. Western blot assays were performed to assess changes in the expression of osteogenic-related markers and the phosphorylation of PI3K and AKT. Finally, we established an ovariectomized (OVX)-induced osteoporosis mouse model and administered curculigoside to explore the effects of curculigoside in preventing bone loss in vivo. Results The CCK-8 assay indicated that curculigoside did not induce cytotoxicity at a concentration of 5 μmol/L after 48 h. The ALP and ARS results revealed that the induced group had higher ALP activity and calcium deposition than the control group. Moreover, the curculigoside group exhibited increased biomineralization, ALP activity, and ARS staining compared to the induced and control groups, and these effects were partially inhibited by LY294002. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis indicated that the target genes of curculigoside were mainly involved in the PI3K-Akt signaling pathway. PCR and western blot analysis showed that the expression of RUNX2, ALP, and Osterix was upregulated in curculigoside-treated ADSCs, but this effect was partially reversed by the PI3K inhibitor LY294002. Moreover, the curculigoside-treated group exhibited significantly increased phosphorylation of AKT to P-AKT compared with the osteogenic induction group. After treatment with curculigoside, the mice had a higher bone volume than the OVX mice, suggesting partial protection from cancellous bone loss. In addition, when LY294002 was added, the protective effects of curculigoside could be neutralized. Conclusions Curculigoside could induce the osteogenic differentiation of ADSCs and prevent bone loss in an OVX model through the PI3K/Akt signaling pathway.

scholarly journals Salidroside promoted osteogenic differentiation of adipose-derived stromal cells through Wnt/β-catenin signaling pathway

2021 ◽  
Vol 16 (1) ◽  
Author(s):  
Xiao-hua Li ◽  
Fu-ling Chen ◽  
Hong-lin Shen
Keyword(s):  
Western Blot ◽  
Osteogenic Differentiation ◽  
Signaling Pathway ◽  
Differentially Expressed Genes ◽  
Stromal Cells ◽  
Differentially Expressed ◽  
Calcium Deposition ◽  
Western Blot Assay ◽  
Adipose Derived Stromal Cells ◽  
Interfering Rna

Abstract Background Bone disease causes short-term or long-term physical pain and disability. It is necessary to explore new drug for bone-related disease. This study aimed to explore the role and mechanism of Salidroside in promoting osteogenic differentiation of adipose-derived stromal cells (ADSCs). Methods ADSCs were isolated and treated with different dose of Salidroside. Cell count kit-8 (CCK-8) assay was performed to assess the cell viability of ADSCs. Then, ALP and ARS staining were conducted to assess the early and late osteogenic capacity of ADSCs, respectively. Then, differentially expressed genes were obtained by R software. Then, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of the differentially expressed genes were further analyzed. The expression of OCN, COL1A1, RUNX2, WNT3A, and β-catenin were measured by real-time PCR and Western blot analysis. Last, β-catenin was silenced by small interfering RNA. Results Salidroside significantly increased the ADSCs viability at a dose-response manner. Moreover, Salidroside enhanced osteogenic capacity of ADSCs, which are identified by enhanced ALP activity and calcium deposition. A total of 543 differentially expressed genes were identified between normal and Salidroside-treated ADSCs. Among these differentially expressed genes, 345 genes were upregulated and 198 genes were downregulated. Differentially expressed genes enriched in the Wnt/β-catenin signaling pathway. Western blot assay indicated that Salidroside enhanced the WNT3A and β-catenin expression. Silencing β-catenin partially reversed the promotion effects of Salidroside. PCR and Western blot results further confirmed these results. Conclusion Salidroside promoted osteogenic differentiation of ADSCs through Wnt/β-catenin signaling pathway.

scholarly journals Astragaloside positively regulated osteogenic differentiation of pre-osteoblast MC3T3-E1 through PI3K/Akt signaling pathway

2021 ◽  
Vol 16 (1) ◽  
Author(s):  
Wei Bing Jing ◽  
Hongjuan Ji ◽  
Rui Jiang ◽  
Jinlong Wang
Keyword(s):  
Western Blot ◽  
Osteogenic Differentiation ◽  
Real Time ◽  
Signaling Pathway ◽  
Real Time Pcr ◽  
Akt Signaling ◽  
Calcium Deposition ◽  
Control Group ◽  
Akt Signaling Pathway ◽  
Western Blot Assay

Abstract Background Osteoporosis is a widespread chronic disease characterized by low bone density. There is currently no gold standard treatment for osteoporosis. The aim of this study was to explore the role and mechanism of Astragaloside on osteogenic differentiation of MC3T3-E1 cells. Methods MC3T3-E1 cells were divided into control and different dose of Astragaloside (10, 20, 40, 50, and 60 μg/ml). Then, ALP and ARS staining were performed to identify the effects of Astragaloside for early and late osteogenic capacity of MC3T3-E1 cells, respectively. Real-time PCR and western blot were performed to assess the ALP, OCN, and OSX expression. PI3K/Akt signaling pathway molecules were then assessed by Western blot. Finally, PI3K inhibitor, LY294002, was implemented to assess the mechanism of Astragaloside in promoting osteogenic differentiation of MC3T3-E1 cells. Results Astragaloside significantly increased the cell viability than the control group. Moreover, Astragaloside enhanced the ALP activity and calcium deposition than the control groups. Compared with the control group, Astragaloside increased the ALP, OCN, and OSX expression in a dose-response manner. Western blot assay further confirmed the real-time PCR results. Astragaloside could significantly increase the p-PI3K and p-Akt expression than the control group. LY294002 partially reversed the promotion effects of Astragaloside on osteogenic differentiation of MC3T3-E1 cells. LY294002 partially reversed the promotion effects of Astragaloside on ALP, OCN, and OSX of MC3T3-E1 cells. Conclusion The present study suggested that Astragaloside promoted osteogenic differentiation of MC3T3-E1 cells through regulating PI3K/Akt signaling pathway.

Abstract P098: Inhibitory Role of Notch1 in Calcific Aortic Valve Disease Is Mediated by Sox9

2011 ◽  
Vol 109 (suppl_1) ◽  
Author(s):  
Chetan P Hans ◽  
Asha Acharya ◽  
Sara N Koenig ◽  
Haley A Nichols ◽  
Cristi L Galindo ◽  
...  
Keyword(s):  
Aortic Valve ◽  
Notch Signaling ◽  
Signaling Pathway ◽  
Aortic Valve Disease ◽  
Notch Signaling Pathway ◽  
Valve Disease ◽  
Calcium Deposition ◽  
Aortic Valve Calcification ◽  
Calcific Aortic Valve Disease ◽  
Valve Calcification

Introduction: Aortic valve calcification is the most common form of valvular heart disease; however the mechanism(s) underlying calcific aortic valve disease (CAVD) are unknown. NOTCH1 mutations are associated with aortic valve malformations and adult-onset calcification in families with inherited disease. The Notch signaling pathway is critical for multiple cell differentiation processes, but its role in the development of CAVD is not well understood. Objective: To investigate the molecular changes associated with the calcification of aortic valve that occurs with inhibition of Notch signaling. Methods and Results: The expression of Notch signaling pathway members was validated in the aortic valve cusps from adult mice, and examination of diseased human aortic valves revealed decreased expression of NOTCH1 in areas of calcium deposition. To identify downstream mediators of Notch1 signaling, we examined gene expression changes that occur with chemical inhibition of Notch signaling in rat aortic valve interstitial cells (AVICs). We found significant downregulation of many cartilage-specific genes that constitute the valve extracellular matrix (ECM). Analysis of these cartilage-specific genes demonstrated that several were transcriptional targets of Sox9, a master regulator of chondrogenesis, which has been previously shown to be essential for proper valve development and maintenance. Utilizing an in vitro porcine aortic valve calcification model system, inhibition of Notch activity resulted in accelerated calcification while stimulation of Notch signaling attenuated the calcific process. Finally, utilizing transfection studies, addition of Sox9 was able to prevent the calcification of porcine AVICs that occurs with Notch inhibition. Conclusions: Loss of Notch signaling contributes to aortic valve calcification by a Sox9-dependent mechanism. Further elucidation of the Notch1-Sox9 molecular pathway and its role in the maintenance of the ECM will lead to an improved mechanistic understanding of aortic valve calcification and development of novel therapeutic strategies for CAVD.

scholarly journals Concentrated Growth Factors (CGF) Induce Osteogenic Differentiation in Human Bone Marrow Stem Cells

Biology ◽  
2020 ◽  
Vol 9 (11) ◽  
pp. 370
Author(s):  
Alessio Rochira ◽  
Luisa Siculella ◽  
Fabrizio Damiano ◽  
Andrea Palermo ◽  
Franco Ferrante ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Osteogenic Differentiation ◽  
Bone Marrow Stem Cells ◽  
Protein Quantification ◽  
Human Bone ◽  
Human Bone Marrow ◽  
Osteogenic Medium ◽  
Alizarin Red ◽  
Matrix Mineralization

Bone regeneration is a complex process regulated by several factors that control overlapping biological processes, coordinating interactions among distinct cell populations. There is a great interest in identifying new strategies for inducing osteogenesis in a safe and efficient manner. Concentrated Growth Factor (CGF) is an autologous blood derived product obtained by centrifugation of venous blood following the procedure set on the Silfradent device. In this study the effects of CGF on osteogenic differentiation of human Bone Marrow Stem Cells (hBMSC) in vitro have been investigated; hBMSC were cultured with CGF or osteogenic medium, for 21 days. The osteogenic differentiation was evaluated measuring alkaline phosphatase (ALP) enzyme activity, matrix mineralization by alizarin red staining and through mRNA and protein quantification of osteogenic differentiation markers by Real-time PCR and Western blotting, respectively. The treatment with CGF stimulated ALP activity and promoted matrix mineralization compared to control and seems to be more effective than osteogenic medium. Also, hBMSC lost mesenchymal markers and showed other osteogenic features. Our study showed for the first time that CGF alone is able to induce osteogenic differentiation in hBMSC. The application of CGF on hBMSC osteoinduction might offer new clinical and biotechnological strategies in the tissue regeneration field.

scholarly journals Effect of puerarin on osteogenic differentiation of human periodontal ligament stem cells

2019 ◽  
Vol 48 (4) ◽  
pp. 030006051985164
Author(s):  
Jun Li ◽  
Youjian Peng
Keyword(s):  
Stem Cells ◽  
Osteogenic Differentiation ◽  
Periodontal Ligament ◽  
Calcium Deposition ◽  
Pcr Analysis ◽  
Cell Surface Markers ◽  
Periodontal Ligament Stem Cells ◽  
Alizarin Red ◽  
Alp Activity ◽  
Experimental Group

Objective To investigate the effects of the flavonoid, puerarin, on osteogenic differentiation of human periodontal ligament stem cells (PDLSCs). Methods Human PDLSCs were isolated from patients undergoing orthodontic treatment, and the cell surface markers CD146, CD34, CD45, and STRO-1 were identified by immunofluorescence. Cell proliferation was detected by MTT assay; alkaline phosphatase (ALP) activity was measured, and calcium deposition was detected by alizarin red staining. PCR was then used to detect the distributions of COL-I, OPN, Runx2, and OCN, genes related to osteogenic differentiation. Results Staining was positive for cytokines CD146, CD34, CD45, and STRO-1 in the experimental group; staining was also positive for silk protein, but negative for keratin. After 7 days of culture, exposure to puerarin significantly promoted the level of intracellular ALP; increased puerarin concentration led to increased intracellular ALP. Red mineralized nodules appeared upon exposure to puerarin and the number of nodules was concentration-dependent. PCR analysis revealed that COL-I, OPN, Runx2, and OCN expression levels increased as puerarin concentration increased. Conclusions Exposure to puerarin can promote proliferation and ALP activity in human PDLSCs, thus promoting both molecular and osteogenic differentiation; these findings may provide a theoretical basis for the clinical treatment of periodontal disease with puerarin.

scholarly journals Preventive Effects of Evodiamine on Dexamethasone-Induced Osteoporosis in Zebrafish

2019 ◽  
Vol 2019 ◽  
pp. 1-6 ◽  
Author(s):  
Heng Yin ◽  
Jianwei Wang ◽  
Mao Wu ◽  
Yong Ma ◽  
Shanfu Wang ◽  
...  
Keyword(s):  
Western Blot ◽  
Mapk Pathway ◽  
Bone Mineralization ◽  
Tartrate Resistant Acid Phosphatase ◽  
Mrna Levels ◽  
Rt Pcr ◽  
Alizarin Red ◽  
Calcium Phosphorus ◽  
Commercial Kits ◽  
Preventive Effects

The aim of this study was to investigate the effect of evodiamine (EV) on dexamethasone-induced osteoporosis in zebrafish. Zebrafish larvae were exposed to different concentrations of dexamethasone to obtain the osteoporosis in zebrafish. Calcium, phosphorus, and alizarin red staining determination were performed to evaluate the effects of EV on bone mineralization. Alkaline phosphatase (ALP), hydroxyproline (HP), and tartrate resistant acid phosphatase (TRAP) were also measured by commercial kits. The expression of MMP3-OPN-MAPK pathway in zebrafish was measured by Western blot. RT-PCR was used to determine mRNA levels of MMP3, OPN, and MAPK. EV could significantly increase the content of calcium and phosphorus. The results of alizarin red staining showed that EV could significantly increase the calcium sink of horse fish, increasing the area of bone formation. EV could increase the content of hydroxyproline in zebrafish. EV also increased ALP and TRAP in zebrafish. Western blot and RT-PCR results showed that EV restored the MMP3-OPN-MAPK pathway in zebrafish. In conclusion, we found that EV can alleviate dexamethasone-induced osteoporosis in zebrafish. The mechanism is related to activating MMP3-OPN-MAPK pathway and then activating bone remodeling.

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