Objective To investigate the effect of TonEBP gene on macrophage activity, migration, proliferation and phagocytosis. Methods The Raw264.7 cells transfected with TonEBP -shRNA lentivirus were screened by purinomycin to establish stable cell lines. The interference efficiency was evaluated by real-time PCR and Western blot. The lentivirus infection group with the highest interference efficiency and the empty lentivirus infection group were selected for subsequent experiments. Cells in each group were simultaneously cultured and incubated for 24 h.The cell activity was measured by CCK-8 assay, and the migration ability was observed by transwell migaration assay. Finally, the cell cycle and phagocytosis ability were detected by flow cytometry. Results The TonEBP stable cell line was successfully established and screened out. Compared with the wild type cell group, the interference efficiency of mRNA and protein was over 70% (P<0.05).Downregulation of TonEBP expression significantly reduced cell activity[24 h (0.39±0.02 vs 0.46±0.01),48 h (0.57±0.02 vs 0.63±0.02),72 h (0.71±0.02 vs 1.01±0.11),96 h (0.93±0.06 vs 1.42±0.04),120 h (1.26±0.06 vs 1.56±0.09) ] (P<0.05), red cell migration (88±7.00 vs 356±35.23), proliferation[ (38.73±2.57) ％ vs (50.20±2.29) ％] and phagocytosis [ (2.26±0.10) ％ vs (5.63±0.42) ％] (P<0.05). Conclusions Interference with TonEBP expression can effectively inhibit the activity of macrophages, reduce their migration, proliferation and phagocytosis ability. These results can contribute to further exploration of diseases related to macrophages.
Key words
tonicity-responsive enhancer binding protein /
Raw264.7 /
activity /
migration /
proliferation /
phagocytosis
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References
[1] Moghaddam A S, Mohammadian S, Vazini H, et al. Macrophage plasticity, polarization and function in health and disease[J]. Journal of Cellμlar Physiology, 2018,12(2):451.
[2] Liu C, Choi H, Johnson Z I, et al.Lack of evidence for involvement of TonEBP and hyperosmotic stimμlus in induction of autophagy in the nucleus pμlposus[J]. Sci Rep,2017,7(1):4543.
[3] Na Y R, Je S, Seok S H.Metabolic features of macrophages in inflammatory diseases and cancer[J].Cancer letters,2018,413:46-58.
[4] Balkwill F R, Mantovani A.Cancer-related inflammation: common themes and therapeutic opportunities[J].Semin Cancer Biol,2012,22(1):33-40.
[5] 张仲柏, 李艳春, 张 梅.免疫细胞自噬在动脉粥样硬化中的作用研究进展[J].武警医学,2019,30(10):909-913.
[6] Rucker A J, Rudemiller N P, Crowley S D.Salt hypertension, and immunity[J]. Annu Rev Physiol,2018,80:283-307.
[7] Thomas D, Apovian C.Macrophage functions in lean and obese adipose tissue[J]. Metabolism,2017,72:120-143.
[8] Italiani P, Boraschi D.From Monocytes to M1/M2 Macrophages: Phenotypical vs. Functional Differentiation[J].Frontiers in immunology,2014,5:514.
[9] Laskin D L, Sunil V R, Gardner C R, et al.Macrophages and tissue injury: agents of defense or destruction?[J]. Annu Rev Pharmacol Toxicol,2011,51:267-288.
[10] Katsumoto A, Miranda A S, Butovsky O, et al.Laquinimod attenuates inflammation by modμlating macrophage functions in traumatic brain injury mouse model[J].Journal of neuroinflammation,2018,15(1):26.
[11] 罗悦晨, 周 欣, 姬文婕,等.干扰NaV1.9对RAW264.7细胞增殖、吞噬和迁移的影响[J].细胞与分子免疫学杂志,2013,29(3):225-228.
[12] Groh L, Keating S T, Joosten L A B, et al.Monocyte and macrophage immunometabolism in atherosclerosis[J]. Semin Immunopathol,2018,40(2):203-214.
[13] Tang Y, Pan B, Zhou X, et al.Wip1-dependent mod?lation of macrophage migration and phagocytosis[J].Redox biology,2017,13:665-673.
[14] Coffman T M.Under pressure: the search for the essential mechanisms of hypertension[J].Nature,2011,17(11):1402-1409.
[15] Durantaye M, Piette A B, Rooijen N, et al.Macrophage depletion reduces cell proliferation and extracellμlar matrix accumμlation but increases the ltimate tensile strength of injured Achilles tendons[J]. J Orthop Res,2014,32(2):279-285.
[16] Buxade M, Lunazzi G, Minguillon J, et al.Gene expression induced by Toll-like receptors in macrophages requires the transcription factor NFAT5[J]. J Exp Med,2012,209(2):379-393.
[17] Yoon H J, You S, Yoo S A, et al.NF-AT5 is a critical regμlator of inflammatory arthritis[J]. Arthritis Rheum,2011,63(7):1843-1852.
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