Objective  To investigate the effect of Dexmedetomidine (Dex) on osteosarcoma (OS) cells and its underlying mechanism.

Methods The optimal exposure concentrations of Dex, α2-adrenergic receptor (α2-AR) antagonist, PKA inhibitor, and ERK1/2 inhibitor were determined by MTT assay. Genes positively correlated with ERK1/2 expression in OS samples and potentially mediating its tumor-promoting effect were extracted from the GEPIA database, and their expression levels in OS cells were verified by qRT-PCR and Western blot. After silencing candidate genes with siRNA, MTT assay, flow cytometry, Transwell assay, and colony formation assay were performed to evaluate the role of ERK1/2 pathway genes in the processes by which Dex affects OS cell proliferation, apoptosis, invasion, and colony formation ability.

Results 25 nM Dex was the optimal concentration for promoting OS cell viability. The ERK1/2 inhibitor (Ravoxertinib) significantly antagonized the enhancement of OS cell viability induced by low-concentration Dex, whereas inhibition of α2-AR or PKA exerted weaker effects. Bioinformatics analysis identified that TRIO, PKN2, CKAP5, and NEK4 were highly associated with the oncogenic function of ERK1/2 in OS. Western blot demonstrated that Ravoxertinib blocked the upregulation of TRIO, PKN2, CKAP5, and NEK4 induced by Dex. Silencing TRIO, PKN2, or CKAP5 effectively suppressed Dex-induced increases in OS cell viability, invasion capacity, and colony formation, while attenuating the inhibitory effect of Dex on cell apoptosis.

Conclusion Low-concentration Dex upregulates the expression of TRIO, PKN2, and CKAP5 in an ERK1/2-dependent manner, thereby promoting the malignant biological potential of OS cells.

1. Yang W, Shan Z, Zhou X, et al. Knockdown of lncRNA GHET1 inhibits osteosarcoma cellsproliferation, invasion, migration and EMT in vitro and in vivo[J]. Cancer Biomark, 2018, 23(4): 589-601.

2. Müller DA, Silvan U. On the biomechanical properties of osteosarcoma cells and their environment[J]. Int J Dev Biol, 2019, 63(1-2):1-8.

3. Wang JY, Yang Y, Ma Y, et al. Potential regulatory role of lncRNA-miRNA-mRNA axis in osteosarcoma[J]. Biomed Pharmacother, 2020, 121: 109627.

4. Zimmerman B. Crystal structure of a full-length human tetraspanin reveals a cholesterol-binding pockett[J]. Cell, 2016, 167(4): 1041-1051.

5. Todd EB, Alex C, Odion B, et al. G Douglas Letson. Do surgical margins affect local recurrence and survival in extremity, nonmetastatic, high-grade osteosarcoma?[J]. Clin Orthop Relat Res, 2016, 474(3): 677-683.

6. Mark BD, Michelle JC, Peter R, et al. Osteosarcoma of the spine: prognostic variables for local recurrence and overall survival, a multicenter ambispective study[J]. J Neurosurg Spine, 2016, 25(1): 59-68.

7. Sha J, Zhang H, Zhao Y, et al. Dexmedetomidine attenuates lipopolysaccharide-induced liver oxidative stress and cell apoptosis in rats by increasing GSK-3β/MKP-1/Nrf2 pathway activity via the α2 adrenergic receptor[J]. Toxicol Appl Pharmacol, 2019, 364: 144-152.

8. Meng L, Li L, Lu S, et al. The protective effect of dexmedetomidine on LPS-induced acute lung injury through the HMGB1-mediated TLR4/NF-κB and PI3K/Akt/mTOR pathways[J]. Mol Immunol, 2018, 94: 7-17.

9. Zheng B, Zhang S, Ying Y, et al. Administration of dexmedetomidine inhibited NLRP3 inflammasome and microglial cell activities in hippocampus of traumatic brain injury rats[J]. Biosci Rep, 2018, 38(5): 20180892.

10. Römer L, Wurster S, Savola JM, et al. Identification and characterization of the imidazoline I2b-binding sites in the hamster brown adipose tissue as a study model for imidazoline receptors[J]. Arch Physiol Biochem, 2003, 111(2): 159-166.

11. Shuji K, Hideo H, Naoko S, et al. Bidirectional effects of dexmedetomidine on human platelet functions in vitro[J]. Eur J Pharmacol, 2015, 766: 122-128.

12. Souhayl D, Andrea P, Virginie J, et al. Dexmedetomidine increases hippocampal phosphorylated extracellular signal-regulated protein kinase 1 and 2 content by an alpha 2-adrenoceptor-independent mechanism: evidence for the involvement of imidazoline I1 receptors[J]. Anesthesiology, 2008, 108(3): 457-466.

13. Robert RJ. Targeting ERK1/2 protein-serine/threonine kinases in human cancers[J]. Pharmacol Res, 2019, 142: 151-168.

14. Wen CY, Chun YW, Yue P, et al. Dexmedetomidine alleviates H2O2-induced oxidative stress and cell necroptosis through activating of α2-adrenoceptor in H9C2 cells[J]. Mol Biol Rep, 2020, 47(5): 3629-3639.

15. Li Q, Chen C, Chen X, et al. Dexmedetomidine attenuates renal fibrosis via α2-adrenergic receptor-dependent inhibition of cellular senescence after renal ischemia/reperfusion[J]. Life Sci, 2018, 207: 1-8.

16. Liu Y, Gu X, Liu Y. The effect of dexmedetomidine on biological behavior of osteosarcoma cells through miR-1307 expression[J]. Am J Transl Res, 2021, 13(5): 4876-4883.

17. Robert RJ. Targeting ERK1/2 protein-serine/threonine kinases in human cancers[J]. Pharmacol Res, 2019, 142: 151-168.

18. Andrew MK, James S, Simon JC. ERK1/2 inhibitors: new weapons to inhibit the RAS-regulated RAF-MEK1/2-ERK1/2 pathway[J]. Pharmacol Ther, 2018, 187: 45-60.

19. Victoria CM, Laura BR, Anita W, et al. Distinctive requirement of PKCε in the control of Rho GTPases in epithelial and mesenchymally transformed lung cancer cells[J]. Oncogene, 2019, 38(27): 5396-5412.

20. Jane L, Tracey AM, Robert EM, et al. The expression and prognostic value of the guanine nucleotide exchange factors (GEFs) Trio, Vav1 and TIAM-1 in human breast cancer[J]. Int Semin Surg Oncol, 2008, 5: 23.

21. Pavithra R, Vishalakshi N, Krishna P, et al. Role of protein kinase N2 (PKN2) in cigarette smoke-mediated oncogenic transformation of oral cells[J]. J Cell Commun Signal, 2018, 12(4): 709-721.

22. Marc AS, Petros C, Thomas M, et al. AURKA, DLGAP5, TPX2, KIF11 and CKAP5: five specific mitosis-associated genes correlate with poor prognosis for non-small cell lung cancer patients[J]. Int J Oncol, 2017, 50(2): 365-372.

23. Fielding AB, Lim S, Montgomery K, et al. A critical role of integrin-linked kinase, ch-TOG and TACC3 in centrosome clustering in cancer cells[J]. Oncogene, 2011, 30(5): 521-534.