The protective effect of DNA aptamer on osteonecrosis of the femoral head by alleviating TNF-α-mediated necroptosis via RIP1/RIP3/MLKL pathway
Background: The process of necroptosis mediated by tumor necrosis factor alpha (TNF-α) might play an important role in the onset and development of the osteonecrosis of the femoral head (ONFH). The dysfunctions of bone microvascular endothelial cells (BMECs) have been identified as an important part...
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| Format: | Article |
| Language: | English |
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Elsevier
2022-09-01
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| Series: | Journal of Orthopaedic Translation |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2214031X22000638 |
| _version_ | 1811334192111288320 |
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| author | Xiaoyu Fan Xin Xu Xinjie Wu Runzhi Xia Fuqiang Gao Qingyu Zhang Wei Sun |
| author_facet | Xiaoyu Fan Xin Xu Xinjie Wu Runzhi Xia Fuqiang Gao Qingyu Zhang Wei Sun |
| author_sort | Xiaoyu Fan |
| collection | DOAJ |
| description | Background: The process of necroptosis mediated by tumor necrosis factor alpha (TNF-α) might play an important role in the onset and development of the osteonecrosis of the femoral head (ONFH). The dysfunctions of bone microvascular endothelial cells (BMECs) have been identified as an important part of pathological processes in the steroid-induced ONFH. An aptamer is a single-stranded DNA or RNA oligonucleotide sequence. Previous studies have designed or screened various aptamers that could bind to specific targets or receptors in order to block their effects. Objective: There are two main objectives in this study: 1) to establish a TNF-α -induced ONFH model in human BMECs in vitro, 2) to verify the effects of the TNF-α aptamer (AptTNF-α) on blocking TNF-α activity in the ONFH model. Methods: Clinical samples were collected for Hematoxylin and Eosin (HE) staining, immunohistochemistry and further BMEC isolation. After cell culture and identification, the cell viability of BMECs after incubation with TNF-α was assessed by Cell Counting Kit-8 (CCK8). The necroptosis of BMECs was detected by the TUNEL and Annexin V-FITC/PI staining. The attenuation of TNF-α cytotoxicity by AptTNF-α was evaluated by CCK8 at first. Then, the molecular mechanism was explored by the quantitative real-time polymerase chain reaction and western blotting. Results: The expression level of TNF-α was significantly up-regulated in bone tissues of ONFH patients. The identification of BMECs was verified by the high expressions of CD31 and vWF. Results from CCK8, TUNEL staining and Annexin V-FITC/PI assay demonstrated reduced cell viability and increased necroptosis of BMECs after TNF-α stimulation. Further investigations showed that TNF-α cytotoxicity could be attenuated by the AptTNF-α in a dose-dependent manner. Necroptosis mediated by TNF-α in the ONFH model was regulated by the receptor-interacting protein kinase 1 (RIPK1)/receptor-interacting protein kinase 3 (RIPK3)/mixed lineage kinase domain-like protein (MLKL) signalling pathway. Conclusion: We established a TNF-α-induced ONFH model in human BMECs in vitro. Our study also demonstrated that the AptTNF-α could protect BMECs from necroptosis by inhibiting the RIP1/RIP3/MLKL signalling pathway.The Translational Potential of this Article: The effective protection from cell necroptosis provided by the DNA aptamer demonstrated its translational potential as a new type of TNF-α inhibitor in clinical treatments for patients with ONFH. |
| first_indexed | 2024-04-13T17:04:12Z |
| format | Article |
| id | doaj.art-667fe73ce2d740f688d385ddc15bce5c |
| institution | Directory Open Access Journal |
| issn | 2214-031X |
| language | English |
| last_indexed | 2024-04-13T17:04:12Z |
| publishDate | 2022-09-01 |
| publisher | Elsevier |
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| series | Journal of Orthopaedic Translation |
| spelling | doaj.art-667fe73ce2d740f688d385ddc15bce5c2022-12-22T02:38:31ZengElsevierJournal of Orthopaedic Translation2214-031X2022-09-01364451The protective effect of DNA aptamer on osteonecrosis of the femoral head by alleviating TNF-α-mediated necroptosis via RIP1/RIP3/MLKL pathwayXiaoyu Fan0Xin Xu1Xinjie Wu2Runzhi Xia3Fuqiang Gao4Qingyu Zhang5Wei Sun6Peking University Health Science Center, China-Japan Friendship School of Clinical Medicine, Beijing, 100029, ChinaPeking Union Medical College China-Japan Friendship School of Clinical Medicine, Beijing, 100029, ChinaPeking University Health Science Center, China-Japan Friendship School of Clinical Medicine, Beijing, 100029, ChinaPeking Union Medical College China-Japan Friendship School of Clinical Medicine, Beijing, 100029, ChinaOrthopedics Department, China-Japan Friendship Hospital, Beijing, 100029, ChinaDepartment of Orthopedics, Shandong Provincial Hospital Affiliated to Shandong First Medical University, No.324, Road Jing Wu Wei Qi, Jinan, 250021, Shandong, ChinaPeking University Health Science Center, China-Japan Friendship School of Clinical Medicine, Beijing, 100029, China; Orthopedics Department, China-Japan Friendship Hospital, Beijing, 100029, China; Corresponding author. Peking University Health Science Center, China-Japan Friendship School of Clinical Medicine, Beijing, 100029, China.Background: The process of necroptosis mediated by tumor necrosis factor alpha (TNF-α) might play an important role in the onset and development of the osteonecrosis of the femoral head (ONFH). The dysfunctions of bone microvascular endothelial cells (BMECs) have been identified as an important part of pathological processes in the steroid-induced ONFH. An aptamer is a single-stranded DNA or RNA oligonucleotide sequence. Previous studies have designed or screened various aptamers that could bind to specific targets or receptors in order to block their effects. Objective: There are two main objectives in this study: 1) to establish a TNF-α -induced ONFH model in human BMECs in vitro, 2) to verify the effects of the TNF-α aptamer (AptTNF-α) on blocking TNF-α activity in the ONFH model. Methods: Clinical samples were collected for Hematoxylin and Eosin (HE) staining, immunohistochemistry and further BMEC isolation. After cell culture and identification, the cell viability of BMECs after incubation with TNF-α was assessed by Cell Counting Kit-8 (CCK8). The necroptosis of BMECs was detected by the TUNEL and Annexin V-FITC/PI staining. The attenuation of TNF-α cytotoxicity by AptTNF-α was evaluated by CCK8 at first. Then, the molecular mechanism was explored by the quantitative real-time polymerase chain reaction and western blotting. Results: The expression level of TNF-α was significantly up-regulated in bone tissues of ONFH patients. The identification of BMECs was verified by the high expressions of CD31 and vWF. Results from CCK8, TUNEL staining and Annexin V-FITC/PI assay demonstrated reduced cell viability and increased necroptosis of BMECs after TNF-α stimulation. Further investigations showed that TNF-α cytotoxicity could be attenuated by the AptTNF-α in a dose-dependent manner. Necroptosis mediated by TNF-α in the ONFH model was regulated by the receptor-interacting protein kinase 1 (RIPK1)/receptor-interacting protein kinase 3 (RIPK3)/mixed lineage kinase domain-like protein (MLKL) signalling pathway. Conclusion: We established a TNF-α-induced ONFH model in human BMECs in vitro. Our study also demonstrated that the AptTNF-α could protect BMECs from necroptosis by inhibiting the RIP1/RIP3/MLKL signalling pathway.The Translational Potential of this Article: The effective protection from cell necroptosis provided by the DNA aptamer demonstrated its translational potential as a new type of TNF-α inhibitor in clinical treatments for patients with ONFH.http://www.sciencedirect.com/science/article/pii/S2214031X22000638AptamerBone microvascular endothelial cellNecroptosisOsteonecrosis of the femoral headTNF-α |
| spellingShingle | Xiaoyu Fan Xin Xu Xinjie Wu Runzhi Xia Fuqiang Gao Qingyu Zhang Wei Sun The protective effect of DNA aptamer on osteonecrosis of the femoral head by alleviating TNF-α-mediated necroptosis via RIP1/RIP3/MLKL pathway Journal of Orthopaedic Translation Aptamer Bone microvascular endothelial cell Necroptosis Osteonecrosis of the femoral head TNF-α |
| title | The protective effect of DNA aptamer on osteonecrosis of the femoral head by alleviating TNF-α-mediated necroptosis via RIP1/RIP3/MLKL pathway |
| title_full | The protective effect of DNA aptamer on osteonecrosis of the femoral head by alleviating TNF-α-mediated necroptosis via RIP1/RIP3/MLKL pathway |
| title_fullStr | The protective effect of DNA aptamer on osteonecrosis of the femoral head by alleviating TNF-α-mediated necroptosis via RIP1/RIP3/MLKL pathway |
| title_full_unstemmed | The protective effect of DNA aptamer on osteonecrosis of the femoral head by alleviating TNF-α-mediated necroptosis via RIP1/RIP3/MLKL pathway |
| title_short | The protective effect of DNA aptamer on osteonecrosis of the femoral head by alleviating TNF-α-mediated necroptosis via RIP1/RIP3/MLKL pathway |
| title_sort | protective effect of dna aptamer on osteonecrosis of the femoral head by alleviating tnf α mediated necroptosis via rip1 rip3 mlkl pathway |
| topic | Aptamer Bone microvascular endothelial cell Necroptosis Osteonecrosis of the femoral head TNF-α |
| url | http://www.sciencedirect.com/science/article/pii/S2214031X22000638 |
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