CD90's role in vascularization and healing of rib fractures: insights from Dll4/notch regulation.

IF 4.8 3区 医学 Q2 CELL BIOLOGY
Inflammation Research Pub Date : 2024-12-01 Epub Date: 2024-10-26 DOI:10.1007/s00011-024-01962-w
Lei Wang, Rui Hu, Pei Xu, Pengkai Gao, Bin Mo, Liya Dong, Fengqing Hu
{"title":"CD90's role in vascularization and healing of rib fractures: insights from Dll4/notch regulation.","authors":"Lei Wang, Rui Hu, Pei Xu, Pengkai Gao, Bin Mo, Liya Dong, Fengqing Hu","doi":"10.1007/s00011-024-01962-w","DOIUrl":null,"url":null,"abstract":"<p><strong>Background: </strong>Vascularization after rib fracture is a crucial physiological process that is essential for the repair and healing of the rib. Studies have shown that CD90 plays a critical role in regulating rib fracture healing, but the underlying mechanism of its role has not been fully elucidated.</p><p><strong>Methods: </strong>CD90 adenovirus knockout mice were used to construct a rib injury model. The bone healing was observed by micro-CT. CD31/EMCN immunofluorescence staining was performed on bone tissue to observe the density of H-shaped and L-shaped blood vessels at the site of bone injury. CD31 and EMCN dual-stained single cells from the rib fracture sites were detected by flow cytometry. The periosteal stem cells transfected with CD90 or Notch1 overexpression and silencing vector were co-cultured with osteoblast MC3T3-E1 in osteogenic induction medium. Moreover, bone microvascular endothelial cells were extracted from the rib injury and co-cultured with the periosteal stem cells transfected with CD90. CCK-8 was used to detect cell viability, RT-qPCR and Western blot were used to detect Notch1, Notch2, Notch3, Notch4, CD31, HIF-1α, CD90, RUNX2, OCN and OPN expression. Alkaline phosphatase (ALP) staining and alizarin red staining were used to observe mineralized nodules. Immunofluorescence staining was used to detect the expression of Dll4, Notch, and CD90 in each group of cells. The angiogenesis experiment was conducted to observe cellular vascular formation.</p><p><strong>Results: </strong>Compared with the Adsh-NC group, the bone healing in the Adsh-CD90 group was significantly impaired, with a marked reduction in the number and volume of blood vessels at the rib fracture site, as evidenced by CD31/EMCN immunofluorescence staining, which showed a reduction in the number of H type vessels at the site of bone injury. It was found that CD90 depletion can inhibit the signaling of Dll4/Notch in the rib fracture site. Furthermore, we found that overexpression of Notch1 reverses the impairment of tubule formation in bone microvascular endothelial cells caused by CD90 suppression.r.Dll4 protein reverses the inhibitory effect of CD90 deletion on periosteal stem cells and MC3T3-E1 cell viability and osteogenesis. In the end, we found that overexpression of Notch1 and CD90 can promote angiogenesis of bone microvascular endothelial cells and Notch pathway activation.</p><p><strong>Conclusion: </strong>CD90 can affect vascular formation in mouse rib fractures, and CD90 may be regulated by Dll4/Notch.</p>","PeriodicalId":13550,"journal":{"name":"Inflammation Research","volume":" ","pages":"2263-2277"},"PeriodicalIF":4.8000,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11632021/pdf/","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Inflammation Research","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.1007/s00011-024-01962-w","RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2024/10/26 0:00:00","PubModel":"Epub","JCR":"Q2","JCRName":"CELL BIOLOGY","Score":null,"Total":0}
引用次数: 0

Abstract

Background: Vascularization after rib fracture is a crucial physiological process that is essential for the repair and healing of the rib. Studies have shown that CD90 plays a critical role in regulating rib fracture healing, but the underlying mechanism of its role has not been fully elucidated.

Methods: CD90 adenovirus knockout mice were used to construct a rib injury model. The bone healing was observed by micro-CT. CD31/EMCN immunofluorescence staining was performed on bone tissue to observe the density of H-shaped and L-shaped blood vessels at the site of bone injury. CD31 and EMCN dual-stained single cells from the rib fracture sites were detected by flow cytometry. The periosteal stem cells transfected with CD90 or Notch1 overexpression and silencing vector were co-cultured with osteoblast MC3T3-E1 in osteogenic induction medium. Moreover, bone microvascular endothelial cells were extracted from the rib injury and co-cultured with the periosteal stem cells transfected with CD90. CCK-8 was used to detect cell viability, RT-qPCR and Western blot were used to detect Notch1, Notch2, Notch3, Notch4, CD31, HIF-1α, CD90, RUNX2, OCN and OPN expression. Alkaline phosphatase (ALP) staining and alizarin red staining were used to observe mineralized nodules. Immunofluorescence staining was used to detect the expression of Dll4, Notch, and CD90 in each group of cells. The angiogenesis experiment was conducted to observe cellular vascular formation.

Results: Compared with the Adsh-NC group, the bone healing in the Adsh-CD90 group was significantly impaired, with a marked reduction in the number and volume of blood vessels at the rib fracture site, as evidenced by CD31/EMCN immunofluorescence staining, which showed a reduction in the number of H type vessels at the site of bone injury. It was found that CD90 depletion can inhibit the signaling of Dll4/Notch in the rib fracture site. Furthermore, we found that overexpression of Notch1 reverses the impairment of tubule formation in bone microvascular endothelial cells caused by CD90 suppression.r.Dll4 protein reverses the inhibitory effect of CD90 deletion on periosteal stem cells and MC3T3-E1 cell viability and osteogenesis. In the end, we found that overexpression of Notch1 and CD90 can promote angiogenesis of bone microvascular endothelial cells and Notch pathway activation.

Conclusion: CD90 can affect vascular formation in mouse rib fractures, and CD90 may be regulated by Dll4/Notch.

CD90 在肋骨骨折的血管化和愈合中的作用:Dll4/notch 调节的启示。
背景:肋骨骨折后的血管化是一个关键的生理过程,对肋骨的修复和愈合至关重要。研究表明,CD90 在调节肋骨骨折愈合中起着关键作用,但其作用的内在机制尚未完全阐明:方法:利用 CD90 腺病毒基因敲除小鼠构建肋骨损伤模型。方法:用 CD90 腺病毒基因敲除小鼠构建肋骨损伤模型,通过显微 CT 观察骨愈合情况。对骨组织进行 CD31/EMCN 免疫荧光染色,观察骨损伤部位 H 型和 L 型血管的密度。流式细胞术检测了肋骨骨折部位的 CD31 和 EMCN 双染色单细胞。用CD90或Notch1过表达和沉默载体转染的骨膜干细胞与成骨细胞MC3T3-E1在成骨诱导培养基中共同培养。此外,从肋骨损伤处提取骨微血管内皮细胞,与转染 CD90 的骨膜干细胞共同培养。CCK-8用于检测细胞活力,RT-qPCR和Western blot用于检测Notch1、Notch2、Notch3、Notch4、CD31、HIF-1α、CD90、RUNX2、OCN和OPN的表达。碱性磷酸酶(ALP)染色和茜素红染色用于观察矿化结节。免疫荧光染色用于检测各组细胞中 Dll4、Notch 和 CD90 的表达。进行血管生成实验以观察细胞血管的形成:结果:与 Adsh-NC 组相比,Adsh-CD90 组的骨愈合明显受损,肋骨骨折部位的血管数量和体积明显减少,CD31/EMCN 免疫荧光染色显示骨损伤部位的 H 型血管数量减少。研究发现,CD90耗竭可抑制肋骨骨折部位的Dll4/Notch信号传导。此外,我们还发现,Notch1的过表达能逆转CD90抑制导致的骨微血管内皮细胞小管形成障碍。r.Dll4蛋白能逆转CD90缺失对骨膜干细胞和MC3T3-E1细胞活力和成骨的抑制作用。最后,我们发现过表达 Notch1 和 CD90 可促进骨微血管内皮细胞的血管生成和 Notch 通路的激活:结论:CD90可影响小鼠肋骨骨折的血管形成,CD90可能受Dll4/Notch调控。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
求助全文
约1分钟内获得全文 求助全文
来源期刊
Inflammation Research
Inflammation Research 医学-免疫学
CiteScore
9.90
自引率
1.50%
发文量
134
审稿时长
3-8 weeks
期刊介绍: Inflammation Research (IR) publishes peer-reviewed papers on all aspects of inflammation and related fields including histopathology, immunological mechanisms, gene expression, mediators, experimental models, clinical investigations and the effect of drugs. Related fields are broadly defined and include for instance, allergy and asthma, shock, pain, joint damage, skin disease as well as clinical trials of relevant drugs.
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
确定
请完成安全验证×
copy
已复制链接
快去分享给好友吧!
我知道了
右上角分享
点击右上角分享
0
联系我们:info@booksci.cn Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。 Copyright © 2023 布克学术 All rights reserved.
京ICP备2023020795号-1
ghs 京公网安备 11010802042870号
Book学术文献互助
Book学术文献互助群
群 号:481959085
Book学术官方微信