基于转录组学分析ADORA3在牙周炎中的表达及调控

doi:10.13701/j.cnki.kqyxyj.2023.07.009

摘要/Abstract

摘要： 目的:探讨腺苷A3受体(adenosine A3 receptor, ADORA3)在牙周炎中的表达模式及调控作用。方法:构建牙周炎动物模型,micro-CT和HE染色观察各组小鼠牙槽骨丧失和牙周组织病理形态,并对结扎侧和对照侧牙龈组织进行转录组测序分析Adora3表达差异,RT-qPCR法检测ADORA3在人和小鼠牙龈组织及巨噬细胞中表达情况,转录组测序分析ADORA3与下游靶基因相关性并通过RT-qPCR法验证。结果:小鼠牙周炎模型构建成功,结扎侧牙龈组织中Adora3表达上调。牙周炎患者牙龈组织中ADORA3表达上调,炎症微环境下Adora3表达降低。转录组测序分析显示Adora3表达与下游靶基因抑制蛋白β-2(arrestin beta-2, ARRB2)呈正相关,且Arrb2基因敲除后Adora3表达下调。结论:基于转录组学分析及体外实验验证,ADORA3在不同来源牙龈组织及巨噬细胞中差异表达,且ADORA3表达水平与ARRB2呈正相关。

关键词: 转录组学分析, 牙周炎, 腺苷A3受体, 抑制蛋白β-2

Abstract: Objective: To investigate the expression pattern of adenosine A3 receptor (ADORA3) in periodontitis and its regulatory role. Methods: Animal models of periodontitis were constructed. Alveolar bone loss and the pathological morphology of periodontal tissues were observed by micro-CT and HE staining in each group of mice. The gingival tissues on the ligature-induced and control sides were subjected to transcriptomic sequencing to analyze the Adora3 expression, and then RT-qPCR was used to observe the expression of ADORA3 in human and mice periodontal tissues and macrophages. The correlation of ADORA3 with downstream target genes was analyzed by transcriptomic sequencing and validated by RT-qPCR. Results: Experimental periodontitis model was successfully established in mice, and the expression of Adora3 was upregulated in periodontal tissues on the ligature-induced side. ADORA3 was expressed highly in periodontal tissues from periodontitis, whereas Adora3 was downregulated in the inflammatory microenvironment. Transcriptomic sequencing results showed that Adora3 level was positively correlated with the downstream target gene arrestin beta-2 (ARRB2). And Adora3 level was down-regulated after Arrb2 knockout. Conclusion: Transcriptomic analysis and in vitro experiments verified that ADORA3 was differentially expressed in periodontal tissues and macrophages, which was positively correlation with ARRB2.

Key words: transcriptomic analysis, periodontitis, adenosine A3 receptor, arrestin beta-2

冯梅婷, 鲁嘉韦, 罗礼君. 基于转录组学分析ADORA3在牙周炎中的表达及调控[J]. 口腔医学研究, 2023, 39(7): 613-618.

FENG Meiting, LU Jiawei, LUO Lijun. Expression of ADORA3 in Periodontitis and Its Regulatory Role by Transcriptomic Analysis[J]. Journal of Oral Science Research, 2023, 39(7): 613-618.

参考文献

[1] Slots J. Periodontitis: Facts, fallacies and the future [J]. Periodontol 2000, 2017, 75(1):7-23.
[2] Yang D, Zhou Q, Labroska V, et al. G protein-coupled receptors: Structure-and function-based drug discovery [J]. Signal Transduct Target Ther, 2021, 6(1): 7.
[3] Hauser AS, Attwood MM, Rask-Andersen M, et al. Trends in gpcr drug discovery: New agents, targets and indications [J]. Nat Rev Drug Discov, 2017, 16(12): 829-842.
[4] Mahmod Al-Qattan MN, Mordi MN. Molecular basis of modulating adenosine receptors activities [J]. Curr Pharm Des, 2019, 25(7): 817-831.
[5] Caton JG, Armitage G, Berglundh T, et al. A new classification scheme for periodontal and peri-implant diseases and conditions - introduction and key changes from the 1999 classification [J]. J Clin Periodontol, 2018, 45 Suppl 20: S1-S8.
[6] Graves DT, Alshabab A, Albiero ML, et al. Osteocytes play an important role in experimental periodontitis in healthy and diabetic mice through expression of rankl [J]. J Clin Periodontol, 2018, 45(3): 285-292.
[7] Ahn S, Shenoy SK, Wei H, et al. Differential kinetic and spatial patterns of beta-arrestin and g protein-mediated erk activation by the angiotensin Ⅱ receptor [J]. J Biol Chem, 2004, 279(34): 35518-35525.
[8] Li H, Hu D, Fan H, et al. β-Arrestin 2 negatively regulates Toll-like receptor 4 (TLR4)-triggered inflammatory signaling via targeting p38 MAPK and interleukin 10 [J]. J Biol Chem, 2014, 289(33): 23075-23085.
[9] Tang J, Zhou S, Zhou F, et al. Inhibitory effect of tanshinone iia on inflammatory response in rheumatoid arthritis through regulating β-arrestin 2 [J]. Exp Ther Med, 2019, 17(5): 3299-3306.
[10] Liu Y, Wang GY, Liu SK, et al. β-arrestin2 stimulates interleukin-17 production and expression of CD4⁺ T lymphocytes in a murine asthma model [J]. Iran J Allergy Asthma Immunol, 2011, 10(3): 171-182.
[11] Tao J, Jiang C, Guo P, et al. A novel grk2 inhibitor alleviates experimental arthritis through restraining Th17 cell differentiation [J]. Biomed Pharmacother, 2023, 157: 113997.
[12] Marcenes W, Kassebaum NJ, Bernabé E, et al. Global burden of oral conditions in 1990-2010: A systematic analysis [J]. J Dent Res, 2013, 92(7): 592-597.
[13] Jiao J, Jing W, Si Y, et al. The prevalence and severity of periodontal disease in mainland china: Data from the fourth national oral health survey (2015-2016) [J]. J Clin Periodontol, 2021, 48(2): 168-179.
[14] Yang B, Pang X, Li Z, et al. Immunomodulation in the treatment of periodontitis: Progress and perspectives [J]. Front Immunol, 2021, 12: 781378.
[15] Hajishengallis G, Chavakis T. Local and systemic mechanisms linking periodontal disease and inflammatory comorbidities [J]. Nat Rev Immunol, 2021, 21(7): 426-440.
[16] Pasquini S, Contri C, Borea PA, et al. Adenosine and inflammation: Here, there and everywhere [J]. Int J Mol Sci, 2021, 22(14):7685.
[17] Ren TH, Lv MM, An XM, et al. Activation of adenosine a3 receptor inhibits inflammatory cytokine production in colonic mucosa of patients with ulcerative colitis by down-regulating the nuclear factor-kappa b signaling [J]. J Dig Dis, 2020, 21(1): 38-45.
[18] Lee HS, Chung HJ, Lee HW, et al. Suppression of inflammation response by a novel A₃ adenosine receptor agonist thio-Cl-IB-MECA through inhibition of Akt and NF-κB signaling [J]. Immunobiology, 2011, 216(9):997-1003.
[19] Stemmer SM, Manojlovic NS, Marinca MV, et al. Namodenoson in advanced hepatocellular carcinoma and child-pugh B cirrhosis: Randomized placebo-controlled clinical trial [J]. Cancers (Basel), 2021, 13(2):187.
[20] Fishman P. Drugs targeting the A3 adenosine receptor: Human clinical study data [J]. Molecules, 2022, 27(12):3680.
[21] Spooner R, DeGuzman J, Lee KL, et al. Danger signal adenosine via adenosine 2a receptor stimulates growth of porphyromonas gingivalis in primary gingival epithelial cells [J]. Mol Oral Microbiol, 2014, 29(2): 67-78.
[22] Bitto A, Oteri G, Pisano M, et al. Adenosine receptor stimulation by polynucleotides (pdrn) reduces inflammation in experimental periodontitis [J]. J Clin Periodontol, 2013, 40(1): 26-32.
[23] Sun CX, Wall NR, Angelov N, et al. Changes in mRNA expression of adenosine receptors in human chronic periodontitis [J]. Chin J Dent Res, 2011, 14(2): 113-120.
[24] Ramos-Junior ES, Pedram M, Lee RE, et al. CD73-dependent adenosine dampens interleukin-1β-induced cxcl8 production in gingival fibroblasts: Association with heme oxygenase-1 and adenosine monophosphate-activated protein kinase [J]. J Periodontol, 2020, 91(2): 253-262.
[25] Yin W, Li Z, Jin M, et al. A complex structure of arrestin-2 bound to a g protein-coupled receptor [J]. Cell Res, 2019, 29(12): 971-983.
[26] Storme J, Cannaert A, Van Craenenbroeck K, et al. Molecular dissection of the human A₃ adenosine receptor coupling with β-arrestin2 [J]. Biochem Pharmacol, 2018, 148: 298-307.
[27] Storme J, Tosh DK, Gao ZG, et al. Probing structure-activity relationship in β-arrestin2 recruitment of diversely substituted adenosine derivatives [J]. Biochem Pharmacol, 2018, 158: 103-113.