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The time-dependent expression of FPR2 and ANXA1 in murine deep vein thrombosis model and its relation to thrombus age

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Abstract

Thrombus age determination in fatal venous thromboembolism cases is an important task for forensic pathologists. In this study, we investigated the time-dependent expressions of formyl peptide receptor 2 (FPR2) and Annexin A1 (ANXA1) in a stasis-induced deep vein thrombosis (DVT) murine model, with the aim of obtaining useful information for thrombus age timing. A total of 75 ICR mice were randomly classified into thrombosis group and control group. In thrombosis group, a DVT model was established by ligating the inferior vena cava (IVC) of mice, and thrombosed IVCs were harvested at 1, 3, 5, 7, 10, 14, and 21 days after modeling. In control group, IVCs without thrombosis were taken as control samples. The expressions of FPR2 and ANXA1 during thrombosis were detected using immunohistochemistry and double immunofluorescence staining. Their protein and mRNA levels in the samples were determined by Western blotting and quantitative real-time PCR. The results reveal that FPR2 was predominantly expressed by intrathrombotic neutrophils and macrophages. ANXA1 expression in the thrombi was mainly distributed in neutrophils, endothelial cells of neovessels, and fibroblastic cells. After thrombosis, the expressions of FPR2 and ANXA1 were time-dependently up-regulated. The percentage of FPR2-positive cells and the level of FPR2 protein significantly elevated at 1, 3, 5 and 7 days after IVC ligation as compared to those at 10, 14 and 21 days after ligation (p < 0.05). Moreover, the mRNA level of FPR2 were significantly higher at 5 days than that at the other post-ligation intervals (p < 0.05). Besides, the levels of ANXA1 mRNA and protein peaked at 10 and 14 days after ligation, respectively. A significant increase in the mRNA level of ANXA1 was found at 10 and 14 days as compared with that at the other post-ligation intervals (p < 0.01). Our findings suggest that FPR2 and ANXA1 are promising as useful markers for age estimation of venous thrombi.

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Funding

This study was supported by the Natural Science Foundation of Zhejiang Province in China (LY20H150006) and National Natural Science Foundation of China (81301640).

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  1. Jun-Jie Huang and Jia-ying Zhuo contributed equally to this work.

Authors and Affiliations

  1. Department of Forensic Medicine, Wenzhou Medical University, Higher Education District, Wenzhou, 325035, Zhejiang Province, People’s Republic of China

    Jun-Jie Huang, Jia-ying Zhuo, Qian Wang, Yue Sun, Jia-Xin Qi, Juan-Juan Wu, Yu Zhang, Gang Chen & Yan-Yan Fan

  2. School of Basic Medical Science, Wenzhou Medical University, Higher Education District, Wenzhou, 325035, Zhejiang Province, People’s Republic of China

    Jun-Jie Huang, Jia-ying Zhuo, Qian Wang, Yue Sun, Jia-Xin Qi, Juan-Juan Wu, Yu Zhang, Gang Chen, Peng-Fei Jiang & Yan-Yan Fan

  3. Forensic Center, Wenzhou Medical University, Wenzhou, 325027, Zhejiang Province, People’s Republic of China

    Gang Chen & Yan-Yan Fan

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Yan-Yan Fan and Peng-Fei Jiang designed the research and wrote the manuscript. Jun-Jie Huang and Jia-ying Zhuo conducted the animal experiments, sample collection, Western blot and Real time PCR. Measurement of thrombus size was conducted by Qian Wang, Yue Sun, and Jia-Xin Qi. Immunohistochemistry, morphometrical analysis, and double immunofluorescence staining were performed by Juan-Juan Wu, Qian Wang, Yue Sun, Jia-Xin Qi, and Yu Zhang. Experimental data were analyzed by Gang Chen. All authors read and approved the final manuscript.

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Huang, JJ., Zhuo, Jy., Wang, Q. et al. The time-dependent expression of FPR2 and ANXA1 in murine deep vein thrombosis model and its relation to thrombus age. Forensic Sci Med Pathol (2024). https://doi.org/10.1007/s12024-024-00818-3

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