Single cell analysis reveals inhibition of angiogenesis attenuates the progression of heterotopic ossification in Mkx −/− mice

Junxin Lin; Yuwei Yang; Wenyan Zhou; Chao Dai; Xiao Chen; Yuanhao Xie; Shan Han; Huanhuan Liu; Yejun Hu; Chenqi Tang; Varitsara Bunpetch; Dandan Zhang; Yishan Chen; Xiaohui Zou; Di Chen; Wanlu Liu; Hongwei Ouyang

doi:10.1038/s41413-021-00175-9

Single cell analysis reveals inhibition of angiogenesis attenuates the progression of heterotopic ossification in Mkx −/− mice

Junxin Lin, Yuwei Yang, Wenyan Zhou, Chao Dai, Xiao Chen, Yuanhao Xie, Shan Han, Huanhuan Liu, Yejun Hu, Chenqi Tang, Varitsara Bunpetch, Dandan Zhang, Yishan Chen, Xiaohui Zou, Di Chen, Wanlu Liu, Hongwei Ouyang

Research output: Contribution to journal › Article › peer-review

12 Citations (Scopus)

Abstract

Tendon heterotopic ossification (HO) is characterized by bone formation inside tendon tissue, which severely debilitates people in their daily life. Current therapies fail to promote functional tissue repair largely due to our limited understanding of HO pathogenesis. Here, we investigate the pathological mechanism and propose a potential treatment method for HO. Immunofluorescence assays showed that the Mohawk (MKX) expression level was decreased in human tendon HO tissue, coinciding with spontaneous HO and the upregulated expression of osteochondrogenic and angiogenic genes in the tendons of Mkx^−/− mice. Single-cell RNA sequencing analyses of wild-type and Mkx^−/− tendons identified three cell types and revealed the excessive activation of osteochondrogenic genes during the tenogenesis of Mkx^−/− tendon cells. Single-cell analysis revealed that the gene expression program of angiogenesis, which is strongly associated with bone formation, was activated in all cell types during HO. Moreover, inhibition of angiogenesis by the small-molecule inhibitor BIBF1120 attenuated bone formation and angiogenesis in the Achilles tendons of both Mkx mutant mice and a rat traumatic model of HO. These findings provide new insights into the cellular mechanisms of tendon HO and highlight the inhibition of angiogenesis with BIBF1120 as a potential treatment strategy for HO.

Original language	English
Article number	4
Number of pages <span style="color:red"p> <font size="1.5"> ✽ </span> </font>	11
Journal	Bone Research
Volume	10
Issue number	4
DOIs	https://doi.org/10.1038/s41413-021-00175-9
Publication status	Published - 7 Jan 2022
Externally published	Yes

Bibliographical note

Funding Information:
We thank Dr. Savio L-Y. Woo and Dr. Mikael Bjorklund for the helpful advice regarding the writing of the manuscript. This work was supported by the National Key R&D Program of China (2017YFA0104900), the National Natural Science Foundation of China (31830029, 81501937 and 81522029), and the Fundamental Research Funds for the Central Universities (K20200099).

Publisher Copyright:
© 2022, The Author(s).

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10.1038/s41413-021-00175-9

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Lin, J., Yang, Y., Zhou, W., Dai, C., Chen, X., Xie, Y., Han, S., Liu, H., Hu, Y., Tang, C., Bunpetch, V., Zhang, D., Chen, Y., Zou, X., Chen, D., Liu, W., & Ouyang, H. (2022). Single cell analysis reveals inhibition of angiogenesis attenuates the progression of heterotopic ossification in Mkx −/− mice. Bone Research, 10(4), [4]. https://doi.org/10.1038/s41413-021-00175-9

Lin, Junxin ; Yang, Yuwei ; Zhou, Wenyan ; Dai, Chao ; Chen, Xiao ; Xie, Yuanhao ; Han, Shan ; Liu, Huanhuan ; Hu, Yejun ; Tang, Chenqi ; Bunpetch, Varitsara ; Zhang, Dandan ; Chen, Yishan ; Zou, Xiaohui ; Chen, Di ; Liu, Wanlu ; Ouyang, Hongwei. / Single cell analysis reveals inhibition of angiogenesis attenuates the progression of heterotopic ossification in Mkx −/− mice. In: Bone Research. 2022 ; Vol. 10, No. 4.

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title = "Single cell analysis reveals inhibition of angiogenesis attenuates the progression of heterotopic ossification in Mkx −/− mice",

abstract = "Tendon heterotopic ossification (HO) is characterized by bone formation inside tendon tissue, which severely debilitates people in their daily life. Current therapies fail to promote functional tissue repair largely due to our limited understanding of HO pathogenesis. Here, we investigate the pathological mechanism and propose a potential treatment method for HO. Immunofluorescence assays showed that the Mohawk (MKX) expression level was decreased in human tendon HO tissue, coinciding with spontaneous HO and the upregulated expression of osteochondrogenic and angiogenic genes in the tendons of Mkx−/− mice. Single-cell RNA sequencing analyses of wild-type and Mkx−/− tendons identified three cell types and revealed the excessive activation of osteochondrogenic genes during the tenogenesis of Mkx−/− tendon cells. Single-cell analysis revealed that the gene expression program of angiogenesis, which is strongly associated with bone formation, was activated in all cell types during HO. Moreover, inhibition of angiogenesis by the small-molecule inhibitor BIBF1120 attenuated bone formation and angiogenesis in the Achilles tendons of both Mkx mutant mice and a rat traumatic model of HO. These findings provide new insights into the cellular mechanisms of tendon HO and highlight the inhibition of angiogenesis with BIBF1120 as a potential treatment strategy for HO.",

author = "Junxin Lin and Yuwei Yang and Wenyan Zhou and Chao Dai and Xiao Chen and Yuanhao Xie and Shan Han and Huanhuan Liu and Yejun Hu and Chenqi Tang and Varitsara Bunpetch and Dandan Zhang and Yishan Chen and Xiaohui Zou and Di Chen and Wanlu Liu and Hongwei Ouyang",

note = "Funding Information: We thank Dr. Savio L-Y. Woo and Dr. Mikael Bjorklund for the helpful advice regarding the writing of the manuscript. This work was supported by the National Key R&D Program of China (2017YFA0104900), the National Natural Science Foundation of China (31830029, 81501937 and 81522029), and the Fundamental Research Funds for the Central Universities (K20200099). Publisher Copyright: {\textcopyright} 2022, The Author(s).",

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doi = "10.1038/s41413-021-00175-9",

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Single cell analysis reveals inhibition of angiogenesis attenuates the progression of heterotopic ossification in Mkx −/− mice. / Lin, Junxin; Yang, Yuwei; Zhou, Wenyan; Dai, Chao; Chen, Xiao; Xie, Yuanhao; Han, Shan; Liu, Huanhuan; Hu, Yejun; Tang, Chenqi; Bunpetch, Varitsara; Zhang, Dandan; Chen, Yishan; Zou, Xiaohui; Chen, Di; Liu, Wanlu; Ouyang, Hongwei.

In: Bone Research, Vol. 10, No. 4, 4, 07.01.2022.

Research output: Contribution to journal › Article › peer-review

TY - JOUR

T1 - Single cell analysis reveals inhibition of angiogenesis attenuates the progression of heterotopic ossification in Mkx −/− mice

AU - Lin, Junxin

AU - Yang, Yuwei

AU - Zhou, Wenyan

AU - Dai, Chao

AU - Chen, Xiao

AU - Xie, Yuanhao

AU - Han, Shan

AU - Liu, Huanhuan

AU - Hu, Yejun

AU - Tang, Chenqi

AU - Bunpetch, Varitsara

AU - Zhang, Dandan

AU - Chen, Yishan

AU - Zou, Xiaohui

AU - Chen, Di

AU - Liu, Wanlu

AU - Ouyang, Hongwei

N1 - Funding Information: We thank Dr. Savio L-Y. Woo and Dr. Mikael Bjorklund for the helpful advice regarding the writing of the manuscript. This work was supported by the National Key R&D Program of China (2017YFA0104900), the National Natural Science Foundation of China (31830029, 81501937 and 81522029), and the Fundamental Research Funds for the Central Universities (K20200099). Publisher Copyright: © 2022, The Author(s).

PY - 2022/1/7

Y1 - 2022/1/7

N2 - Tendon heterotopic ossification (HO) is characterized by bone formation inside tendon tissue, which severely debilitates people in their daily life. Current therapies fail to promote functional tissue repair largely due to our limited understanding of HO pathogenesis. Here, we investigate the pathological mechanism and propose a potential treatment method for HO. Immunofluorescence assays showed that the Mohawk (MKX) expression level was decreased in human tendon HO tissue, coinciding with spontaneous HO and the upregulated expression of osteochondrogenic and angiogenic genes in the tendons of Mkx−/− mice. Single-cell RNA sequencing analyses of wild-type and Mkx−/− tendons identified three cell types and revealed the excessive activation of osteochondrogenic genes during the tenogenesis of Mkx−/− tendon cells. Single-cell analysis revealed that the gene expression program of angiogenesis, which is strongly associated with bone formation, was activated in all cell types during HO. Moreover, inhibition of angiogenesis by the small-molecule inhibitor BIBF1120 attenuated bone formation and angiogenesis in the Achilles tendons of both Mkx mutant mice and a rat traumatic model of HO. These findings provide new insights into the cellular mechanisms of tendon HO and highlight the inhibition of angiogenesis with BIBF1120 as a potential treatment strategy for HO.

AB - Tendon heterotopic ossification (HO) is characterized by bone formation inside tendon tissue, which severely debilitates people in their daily life. Current therapies fail to promote functional tissue repair largely due to our limited understanding of HO pathogenesis. Here, we investigate the pathological mechanism and propose a potential treatment method for HO. Immunofluorescence assays showed that the Mohawk (MKX) expression level was decreased in human tendon HO tissue, coinciding with spontaneous HO and the upregulated expression of osteochondrogenic and angiogenic genes in the tendons of Mkx−/− mice. Single-cell RNA sequencing analyses of wild-type and Mkx−/− tendons identified three cell types and revealed the excessive activation of osteochondrogenic genes during the tenogenesis of Mkx−/− tendon cells. Single-cell analysis revealed that the gene expression program of angiogenesis, which is strongly associated with bone formation, was activated in all cell types during HO. Moreover, inhibition of angiogenesis by the small-molecule inhibitor BIBF1120 attenuated bone formation and angiogenesis in the Achilles tendons of both Mkx mutant mice and a rat traumatic model of HO. These findings provide new insights into the cellular mechanisms of tendon HO and highlight the inhibition of angiogenesis with BIBF1120 as a potential treatment strategy for HO.

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Single cell analysis reveals inhibition of angiogenesis attenuates the progression of heterotopic ossification in Mkx −/− mice

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