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ROCK1 inhibition improves wound healing in diabetes via RIPK4/AMPK pathway

Acta Pharmacologica Sinica (2024)Cite this article

Abstract

Refractory wounds are a severe complication of diabetes mellitus that often leads to amputation because of the lack of effective treatments and therapeutic targets. The pathogenesis of refractory wounds is complex, involving many types of cells. Rho-associated protein kinase-1 (ROCK1) phosphorylates a series of substrates that trigger downstream signaling pathways, affecting multiple cellular processes, including cell migration, communication, and proliferation. The present study investigated the role of ROCK1 in diabetic wound healing and molecular mechanisms. Our results showed that ROCK1 expression significantly increased in wound granulation tissues in diabetic patients, streptozotocin (STZ)-induced diabetic mice, and db/db diabetic mice. Wound healing and blood perfusion were dose-dependently improved by the ROCK1 inhibitor fasudil in diabetic mice. In endothelial cells, fasudil and ROCK1 siRNA significantly elevated the phosphorylation of adenosine monophosphate-activated protein kinase at Thr172 (pThr172-AMPKα), the activity of endothelial nitric oxide synthase (eNOS), and suppressed the levels of mitochondrial reactive oxygen species (mtROS) and nitrotyrosine formation. Experiments using integrated bioinformatics analysis and coimmunoprecipitation established that ROCK1 inhibited pThr172-AMPKα by binding to receptor-interacting serine/threonine kinase 4 (RIPK4). These results suggest that fasudil accelerated wound repair and improved angiogenesis at least partially through the ROCK1/RIPK4/AMPK pathway. Fasudil may be a potential treatment for refractory wounds in diabetic patients.

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Fig. 1: ROCK1 expression levels in diabetic wounds.
Fig. 2: ROCK1 inhibition promoted diabetic wound healing.
Fig. 3: ROCK1 inhibition reduced ROS and increased NO production.
Fig. 4: ROCK1 inhibition enhanced EC angiogenesis.
Fig. 5: ROCK1 inhibition promoted diabetic wound healing.
Fig. 6: ROCK1 inhibition increased the ratio of pThr172-AMPKα/AMPKα under HG.
Fig. 7: RIPK4 mediated ROCK1-induced regulation of AMPKα.
Fig. 8: Schematic diagram of the possible mechanism contributing to diabetic wound healing regulated by ROCK1 in endothelial cells.

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Acknowledgements

This work was supported by the National Natural Science Foundation of China (No. 92168120, 81974506, and 81673486 to LT, No. 81874318, 81673453, 81473235 and 82073878 to XJL, No. 82204494 to YQD), the Beijing Natural Science Foundation (No. Z200019 and 7172119 to LT, No. 7232092 to YQD), Xinjiang Uygur Autonomous Region Key Research and Development Program (No. 2023B02010 to LT), the “Capital Clinical Diagnosis and Treatment Technology Research and Demonstration Application” project commissioned by the Beijing Municipal Commission of Science and Technology (NO. Z191100006619016 to LZ).

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  1. These authors contributed equally: Tianru Huyan, Lu Fan, Zhong-yuan Zheng

Authors and Affiliations

  1. Department of Pharmacology, School of Basic Medical Sciences, Peking University and Beijing Key Laboratory of Tumor Systems Biology, Peking University, Beijing, 100191, China

    Tianru Huyan, Lu Fan, Zhong-yuan Zheng, Zhen-ru Han, Pin Wu, Qun Ma, Ya-qin Du, Yun-di Shi, Xue-jun Li & Lu Tie

  2. Department of Wound Healing Center and Interventional Radiology and Vascular Surgery, Peking University Third Hospital, Beijing, 100191, China

    Tianru Huyan, Jing-hui Zhao & Long Zhang

  3. School of Medicine & Holistic Integrative Medicine, Nanjing University of Chinese Medicine, Nanjing, 210023, China

    Lu Fan, Zhen-ru Han & Chun-yan Gu

  4. Department of Dermatology, Peking University Third Hospital, Beijing, 100191, China

    Wen-hui Wang

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Contributions

TRHY conceived the idea and designed the study. TRHY, LF, ZYZ performed the experiments and analyzed the data. LF, ZYZ and PW wrote the manuscript. ZRH and QM participated in the laser doppler measurements. YDS helped with the immunohistochemistry. JHZ, YQD, CYG, XJL, WHW helped to draft the manuscript. LT and LZ participated in the design and reviewed the manuscript. All authors read and approved the final manuscript.

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Correspondence to Long Zhang or Lu Tie.

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Huyan, T., Fan, L., Zheng, Zy. et al. ROCK1 inhibition improves wound healing in diabetes via RIPK4/AMPK pathway. Acta Pharmacol Sin (2024). https://doi.org/10.1038/s41401-024-01246-3

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