Liuwei Dihuang Pills Enhance Osteogenic Differentiation in MC3T3-E1 Cells through the Activation of the Wnt/β-Catenin Signaling Pathway
Objective: The therapeutic efficacy and molecular mechanisms of traditional Chinese medicines (TCMs), such as Liuwei Dihuang pills (LWDH pills), in treating osteoporosis (OP) remain an area of active research and interest in modern medicine. This study investigated the mechanistic underpinnings of L...
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| Format: | Article |
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MDPI AG
2024-01-01
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| Series: | Pharmaceuticals |
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| Online Access: | https://www.mdpi.com/1424-8247/17/1/99 |
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| author | Jinlong Zhao Guihong Liang Junzheng Yang Hetao Huang Yaoxing Dou Zhuoxu Gu Jun Liu Lingfeng Zeng Weiyi Yang |
| author_facet | Jinlong Zhao Guihong Liang Junzheng Yang Hetao Huang Yaoxing Dou Zhuoxu Gu Jun Liu Lingfeng Zeng Weiyi Yang |
| author_sort | Jinlong Zhao |
| collection | DOAJ |
| description | Objective: The therapeutic efficacy and molecular mechanisms of traditional Chinese medicines (TCMs), such as Liuwei Dihuang pills (LWDH pills), in treating osteoporosis (OP) remain an area of active research and interest in modern medicine. This study investigated the mechanistic underpinnings of LWDH pills in the treatment of OP based on network pharmacology, bioinformatics, and in vitro experiments. Methods: The active ingredients and targets of LWDH pills were retrieved through the TCMSP database. OP-related targets were identified using the CTD, GeneCards, and DisGeNET databases. The STRING platform was employed to construct a protein–protein interaction (PPI) network, and core targets for LWDH pills in treating OP were identified. The GO functional and KEGG pathway enrichment analyses for potential targets were performed using the R package “clusterProfiler”. A “drug–target” network diagram was created using Cytoscape 3.7.1 software. The viability of MC3T3-E1 cells was evaluated using the CCK-8 method after treatment with various concentrations (1.25%, 2.5%, 5%, and 10%) of LWDH pill-medicated serum for 24, 48, and 72 h. Following a 48 h treatment of MC3T3-E1 cells with LWDH pill-medicated serum, the protein levels of collagen Ⅰ, RUNX2, Wnt3, and β-catenin were quantified using the Western blot analysis, and the activity of alkaline phosphatase (ALP) was measured. Results: A total of 197 putative targets for LWDH pills for OP treatment were pinpointed, from which 20 core targets were singled out, including <i>TP53</i>, <i>JUN</i>, <i>TNF</i>, <i>CTNNB1</i> (β-catenin), and <i>GSK3B</i>. The putative targets were predominantly involved in signaling pathways such as the Wnt signaling pathway, the MAPK signaling pathway, and the PI3K-Akt signaling pathway. The intervention with LWDH pill-medicated serum for 24, 48, and 72 h did not result in any notable alterations in the cell viability of MC3T3-E1 cells relative to the control group (all <i>p</i> > 0.05). Significant upregulation in protein levels of collagen Ⅰ, RUNX2, Wnt3, and β-catenin in MC3T3-E1 cells was observed in response to the treatment with 2.5%, 5%, and 10% of LWDH pill-medicated serum in comparison to that with the 10% rabbit serum group (all <i>p <</i> 0.05). Furthermore, the intervention with LWDH pill-medicated serum resulted in the formation of red calcified nodules in MC3T3-E1 cells, as indicated by ARS staining. Conclusions: LWDH pills may upregulate the Wnt/β-catenin signaling pathway to elevate the expression of osteogenic differentiation proteins, including collagen Ⅰ and RUNX2, and to increase the ALP activity in MC3T3-E1 cells for the treatment of OP. |
| first_indexed | 2024-03-08T10:38:19Z |
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| language | English |
| last_indexed | 2024-03-08T10:38:19Z |
| publishDate | 2024-01-01 |
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| spelling | doaj.art-51516eabbe0c4fd8a3e6f1b891f0c7502024-01-26T18:06:03ZengMDPI AGPharmaceuticals1424-82472024-01-011719910.3390/ph17010099Liuwei Dihuang Pills Enhance Osteogenic Differentiation in MC3T3-E1 Cells through the Activation of the Wnt/β-Catenin Signaling PathwayJinlong Zhao0Guihong Liang1Junzheng Yang2Hetao Huang3Yaoxing Dou4Zhuoxu Gu5Jun Liu6Lingfeng Zeng7Weiyi Yang8State Key Laboratory of Traditional Chinese Medicine Syndrome, The Second Clinical College of Guangzhou University of Chinese Medicine, Guangzhou 510405, ChinaState Key Laboratory of Traditional Chinese Medicine Syndrome, The Second Clinical College of Guangzhou University of Chinese Medicine, Guangzhou 510405, ChinaThe Fifth Clinical College, Guangzhou University of Chinese Medicine, Guangzhou 510405, ChinaGuangdong Provincial Key Laboratory of Chinese Medicine for Prevention and Treatment of Refractory Chronic Diseases, The Second Affiliated Hospital of Guangzhou University of Chinese Medicine (Guangdong Provincial Hospital of Chinese Medicine), Guangzhou 510120, ChinaState Key Laboratory of Traditional Chinese Medicine Syndrome, The Second Clinical College of Guangzhou University of Chinese Medicine, Guangzhou 510405, ChinaState Key Laboratory of Traditional Chinese Medicine Syndrome, The Second Clinical College of Guangzhou University of Chinese Medicine, Guangzhou 510405, ChinaThe Research Team on Bone and Joint Degeneration and Injury, Guangdong Provincial Academy of Chinese Medical Sciences, Guangzhou 510120, ChinaState Key Laboratory of Traditional Chinese Medicine Syndrome, The Second Clinical College of Guangzhou University of Chinese Medicine, Guangzhou 510405, ChinaGuangdong Provincial Key Laboratory of Chinese Medicine for Prevention and Treatment of Refractory Chronic Diseases, The Second Affiliated Hospital of Guangzhou University of Chinese Medicine (Guangdong Provincial Hospital of Chinese Medicine), Guangzhou 510120, ChinaObjective: The therapeutic efficacy and molecular mechanisms of traditional Chinese medicines (TCMs), such as Liuwei Dihuang pills (LWDH pills), in treating osteoporosis (OP) remain an area of active research and interest in modern medicine. This study investigated the mechanistic underpinnings of LWDH pills in the treatment of OP based on network pharmacology, bioinformatics, and in vitro experiments. Methods: The active ingredients and targets of LWDH pills were retrieved through the TCMSP database. OP-related targets were identified using the CTD, GeneCards, and DisGeNET databases. The STRING platform was employed to construct a protein–protein interaction (PPI) network, and core targets for LWDH pills in treating OP were identified. The GO functional and KEGG pathway enrichment analyses for potential targets were performed using the R package “clusterProfiler”. A “drug–target” network diagram was created using Cytoscape 3.7.1 software. The viability of MC3T3-E1 cells was evaluated using the CCK-8 method after treatment with various concentrations (1.25%, 2.5%, 5%, and 10%) of LWDH pill-medicated serum for 24, 48, and 72 h. Following a 48 h treatment of MC3T3-E1 cells with LWDH pill-medicated serum, the protein levels of collagen Ⅰ, RUNX2, Wnt3, and β-catenin were quantified using the Western blot analysis, and the activity of alkaline phosphatase (ALP) was measured. Results: A total of 197 putative targets for LWDH pills for OP treatment were pinpointed, from which 20 core targets were singled out, including <i>TP53</i>, <i>JUN</i>, <i>TNF</i>, <i>CTNNB1</i> (β-catenin), and <i>GSK3B</i>. The putative targets were predominantly involved in signaling pathways such as the Wnt signaling pathway, the MAPK signaling pathway, and the PI3K-Akt signaling pathway. The intervention with LWDH pill-medicated serum for 24, 48, and 72 h did not result in any notable alterations in the cell viability of MC3T3-E1 cells relative to the control group (all <i>p</i> > 0.05). Significant upregulation in protein levels of collagen Ⅰ, RUNX2, Wnt3, and β-catenin in MC3T3-E1 cells was observed in response to the treatment with 2.5%, 5%, and 10% of LWDH pill-medicated serum in comparison to that with the 10% rabbit serum group (all <i>p <</i> 0.05). Furthermore, the intervention with LWDH pill-medicated serum resulted in the formation of red calcified nodules in MC3T3-E1 cells, as indicated by ARS staining. Conclusions: LWDH pills may upregulate the Wnt/β-catenin signaling pathway to elevate the expression of osteogenic differentiation proteins, including collagen Ⅰ and RUNX2, and to increase the ALP activity in MC3T3-E1 cells for the treatment of OP.https://www.mdpi.com/1424-8247/17/1/99Liuwei Dihuang pillsosteogenicosteoporosisMC3T3-E1 cellsWnt/β-catenin signaling pathway |
| spellingShingle | Jinlong Zhao Guihong Liang Junzheng Yang Hetao Huang Yaoxing Dou Zhuoxu Gu Jun Liu Lingfeng Zeng Weiyi Yang Liuwei Dihuang Pills Enhance Osteogenic Differentiation in MC3T3-E1 Cells through the Activation of the Wnt/β-Catenin Signaling Pathway Pharmaceuticals Liuwei Dihuang pills osteogenic osteoporosis MC3T3-E1 cells Wnt/β-catenin signaling pathway |
| title | Liuwei Dihuang Pills Enhance Osteogenic Differentiation in MC3T3-E1 Cells through the Activation of the Wnt/β-Catenin Signaling Pathway |
| title_full | Liuwei Dihuang Pills Enhance Osteogenic Differentiation in MC3T3-E1 Cells through the Activation of the Wnt/β-Catenin Signaling Pathway |
| title_fullStr | Liuwei Dihuang Pills Enhance Osteogenic Differentiation in MC3T3-E1 Cells through the Activation of the Wnt/β-Catenin Signaling Pathway |
| title_full_unstemmed | Liuwei Dihuang Pills Enhance Osteogenic Differentiation in MC3T3-E1 Cells through the Activation of the Wnt/β-Catenin Signaling Pathway |
| title_short | Liuwei Dihuang Pills Enhance Osteogenic Differentiation in MC3T3-E1 Cells through the Activation of the Wnt/β-Catenin Signaling Pathway |
| title_sort | liuwei dihuang pills enhance osteogenic differentiation in mc3t3 e1 cells through the activation of the wnt β catenin signaling pathway |
| topic | Liuwei Dihuang pills osteogenic osteoporosis MC3T3-E1 cells Wnt/β-catenin signaling pathway |
| url | https://www.mdpi.com/1424-8247/17/1/99 |
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