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摘要: 目的 探讨褪黑素受体1A （MT1）和褪黑素受体1B （MT2） 敲除对老化相关心房颤动（AF）的影响。方法 将雄性C57BL/6J小鼠分为6组： 8周对照组：野生型，6只，8周龄，体重21～23 g；8周MT1组： MT1敲除，6只，8周龄，体重21～23 g；8周MT2组： MT2敲除，6只，8周龄，体重21～23 g；12个月对照组：野生型，6只，12月龄，体重24～28 g；12个月MT1组： MT1敲除，6只，12月龄，体重24～28 g；12个月MT2组： MT2敲除，6只，12月龄，体重24～28 g。通过超声检测6组小鼠的左心房（LA）内径，电刺激评估小鼠AF诱导率及时间。通过HE染色、Masson染色、DHE染色、TUNEL凋亡染色、免疫荧光染色以及免疫组化染色评估小鼠心房炎症、纤维化、氧化应激和细胞凋亡水平。结果 超声心动图显示，与8周MT1组比较，12个月 MT1组小鼠的LA内径明显增大；与12个月对照组比较，12个月MT1组小鼠的LA内径增大，差异均有统计学意义（P＜0.05）。电刺激结果显示，与8周MT1组比较，12个月 MT1组小鼠的AF诱导率与持续时间明显提高，差异有统计学意义（P＜0.05）。炎症指标： iNOS免疫组织化学染色结果显示，与8周各组比较，12个月对照组、MT1及MT2组小鼠心房iNOS的表达均增加；与12个月对照组比较，12个月 MT1、MT2组小鼠心房iNOS的表达增加，差异均有统计学意义（P＜0.05）。纤维化水平指标：Masson染色显示，与8周各组比较，12个月对照组、MT1及MT2组小鼠心房纤维化水平均增加；与12个月对照组比较，12个月 MT1、MT2组小鼠的心房纤维化水平增加，差异均有统计学意义（P＜0.05）。免疫荧光染色结果显示，与8周各组比较，12个月对照组、MT1及MT2组小鼠心房Ⅲ型胶原蛋白（Collagen Ⅲ）的表达水平均增加；与12个月对照组比较，12个月 MT1、MT2组小鼠心房Collagen Ⅲ的表达水平增加，差异均有统计学意义（P＜0.05）。α-SMA免疫组织化学染色结果显示，与8周各组比较，12个月对照组、MT1及MT2组小鼠心房α-SMA的表达均增加；与12个月对照组比较，12个月 MT1、MT2组小鼠心房α-SMA的表达增加，差异均有统计学意义（P＜0.05）。氧化应激指标：DHE染色结果显示，与8周各组比较，12个月对照组、MT1及MT2组小鼠心房氧化应激水平均增加；与12个月对照组比较，12个月 MT1、MT2组小鼠心房氧化应激水平增加，差异均有统计学意义（P＜0.05）。γ-H2AX免疫荧光染色结果显示，与8周MT1、MT2组比较，12个月 MT1、MT2组小鼠心房中γ-H2AX的表达水平明显增加；与12个月对照组比较，12个月MT1、MT2组小鼠心房中γ-H2AX的表达水平明显增加，差异均有统计学意义（P＜0.05）。凋亡指标：TUNEL结果显示，与8周各组比较，12个月对照组、MT1及MT2组小鼠心房肌细胞凋亡细胞数均增加；与12个月对照组比较，12个月MT1、MT2组小鼠的心房肌细胞凋亡细胞数均增加，差异均有统计学意义（P＜0.05）。p16免疫组织化学结果显示，与8周各组比较，12个月对照组、MT1及MT2组小鼠心房p16表达量均增加；与12个月对照组比较，12个月MT1组小鼠心房p16表达水平增高，差异均有统计学意义（P＜0.05）。结论 MT1和MT2，尤其是MT1，是影响小鼠心房老化进程的主要因素之一，其干预治疗可能延缓AF的发生和发展。Abstract: Objective To investigate the effects of melatonin receptor 1A (MT1) and melatonin receptor 1B (MT2) knockout on age-related atrial fibrillation (AF).Methods Male C57BL/6J mice were divided into 6 groups: 8-week control group (n=6, wild type, 8 weeks old, weight 21-23 g); 8-week MT1 group (n=6, MT1 knocked out, 8 weeks old, weight 21-23 g); 8-week MT2 group (n=6, MT2 knocked out, 8 weeks old, weight 21-23 g); 12-month control group (n=6, wild type, 12 months old, weight 24-28 g); 12-month MT1 group (n=6, MT1 knocked out, 12 months old, weight 24-28 g); 12-month MT2 group (n=6, MT2 knocked out, 12 months old, weight 24-28 g). Left atrial (LA) diameter was measured by echocardiography and AF induction rate and during were evaluated by electrical stimulation. Atrial fibrosis, inflammation, oxidative stress and apoptosis were evaluated by HE, Masson, DHE, TUNEL apoptosis, immunofluorescence and immunohistochemical stains.Results Compared with 8-week MT1 group, the LA diameter in 12-month MT1 group increased significantly. Compared with 12-month control group, the LA diameter in 12-month MT1 group also increased significantly (P＜0.05). Electrical stimulation demonstrated that the AF induction rate and duration in 12-month MT1 group were significantly increased compared with 8-week MT1 group (P＜0.05). Inflammatory indicator: iNOS immunohistochemical staining showed increased iNOS expression in the atrial tissue of 12-month control, MT1 and MT2 groups compared with 8-week groups. Compared to 12-month control group, 12-month MT1 and MT2 groups exhibited significantly higher iNOS expression (P＜0.05). Indicators of fibrosis level: Masson staining revealed that the atrial fibrosis levels increased in 12-month control, MT1 and MT2 groups compared with 8-week groups. The 12-month MT1 and MT2 groups showed significantly higher fibrosis levels than the 12-month control group (P＜0.05). Type Ⅲ collagen expression increased in the 12-month control, MT1, and MT2 groups compared to the 8-week groups. The 12-month MT1 and MT2 groups had significantly higher levels than the 12-month control group (P＜0.05). α-SMA immunohistochemical staining showed that the expression of α-SMA in the atrial tissue of 12-month control, MT1 group and MT2 groups was increased compared with 8-week groups. The 12-month MT1 and MT2 groups had significantly higher levels than the 12-month control group (P＜0.05). Indicators of oxidative stress: DHE staining showed that the oxidative stress levels were elevated in 12-month control, MT1, and MT2 groups compared to 8-week groups. The 12-month MT1 and MT2 groups showed significantly higher oxidative stress levels than the 12-month control group (P＜0.05). γ-H2AX immunofluorescence staining showed that compared with 8-week MT1 and MT2 groups, the expression level of γ-H2AX in the atrial tissue of 12-month MT1 and MT2 groups was significantly increased. Compared with 12-month control group, the expression level of γ-H2AX in 12-month MT1/2 groups was significantly increased (P＜0.05). Indicators of apoptosis: TUNEL assay showed that compared with 8-week groups, the number of apoptotic cells increased in 12-month control, MT1 and MT2 groups. Compared with 12-month control group, the number of apoptotic cells of atrial myocytes in 12-month MT1 and MT2 groups increased significantly (P＜0.05). p16 immunohistochemical stain showed that compared with 8-week groups, the expression of p16 in the atria of 12-month control, MT1 and MT2 groups increased. Compared with 12-month control group, the expression level of p16 in 12-month MT1 group was increased significantly (P＜0.05).Conclusion MT1 and MT2, especially MT1, are key factors affecting atrial aging in mice, and targeted interventions may delay the progression of AF.
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Keywords:
- atrial fibrillation /
- melatonin receptor 1A /
- melatonin receptor 1B
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图 2 电刺激诱导各组小鼠AF成功率及持续时间
Figure 2. Comparison of AF rates and duration induced by electrical stimulation among each group of mice
图 4 各组小鼠心房纤维化水平比较
Figure 4. Comparison of atrial fibrosis levels among each group of mice
图 5 各组小鼠心房氧化应激水平比较
Figure 5. Comparison of atrial oxidative stress among each group of mice
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[1] Liu CB, Zeng J, Wu J, et al. Identification and validation of key genes associated with atrial fibrillation in the elderly[J]. Front Cardiovasc Med, 2023, 10: 1118686. DOI: 10.3389/fcvm.2023.1118686.
[2] Gao P, Gao XY, Xie BX, et al. Aging and atrial fibrillation: a vicious circle[J]. Int J Cardiol, 2024, 395: 131445. DOI: 10.1016/j.ijcard.2023.131445.
[3] Koh AS, Siau A, Gao F, et al. Left atrial phasic function in older adults is associated with fibrotic and low-grade inflammatory pathways[J]. Gerontology, 2023, 69(1): 47-56. DOI: 10.1159/000522632.
[4] Dong YX, Cao XL, Huang JS, et al. Melatonin inhibits fibroblast cell functions and hypertrophic scar formation by enhancing autophagy through the MT2 receptor-inhibited PI3K/Akt/mTOR signaling[J]. Biochim Biophys Acta Mol Basis Dis, 2024, 1870(1): 166887. DOI: 10.1016/j.bbadis.2023.166887.
[5] Nasini S, Tidei S, Shkodra A, et al. Age-related effects of exogenous melatonin on anxiety-like behavior in C57/B6J mice[J]. Biomedicines, 2023, 11(6): 1705. DOI: 10.3390/biomedicines11061705.
[6] Liu JB, Clough SJ, Hutchinson AJ, et al. MT1 and MT2 melatonin receptors: a therapeutic perspective[J]. Annu Rev Pharmacol Toxicol, 2016, 56: 361-383. DOI: 10.1146/annurev-pharmtox-010814-124742.
[7] Durkina AV, Szeiffova Bacova B, Bernikova OG, et al. Blockade of melatonin receptors abolishes its antiarrhythmic effect and slows ventricular conduction in rat hearts[J]. Int J Mol Sci, 2023, 24(15): 11931. DOI: 10.3390/ijms241511931.
[8] Yu LM, Dong X, Zhao JK, et al. Activation of PKG-CREB-KLF15 by melatonin attenuates Angiotensin Ⅱ-induced vulnerability to atrial fibrillation via enhancing branched-chain amino acids catabolism[J]. Free Radic Biol Med, 2022, 178: 202-214. DOI: 10.1016/j.freeradbiomed.2021.11.043.
[9] Xie X, Shen TT, Bi HL, et al. Melatonin inhibits angiotensin Ⅱ-induced atrial fibrillation through preventing degradation of Ang Ⅱ Type Ⅰ Receptor-Associated Protein (ATRAP)[J]. Biochem Pharmacol, 2022, 202: 115146. DOI: 10.1016/j.bcp.2022.115146.
[10] Rose RA, Howlett SE. Preclinical studies on the effects of frailty in the aging heart[J]. Can J Cardiol, 2024, 40(8): 1379-1393. DOI: 10.1016/j.cjca.2024.03.003.
[11] Borgoni S, Kudryashova KS, Burka K, et al. Targeting immune dysfunction in aging[J]. Ageing Res Rev, 2021, 70: 101410. DOI: 10.1016/j.arr.2021.101410.
[12] Ma YG, Mouton AJ, Lindsey ML. Cardiac macrophage biology in the steady-state heart, the aging heart, and following myocardial infarction[J]. Transl Res, 2018, 191: 15-28. DOI: 10.1016/j.trsl.2017.10.001.
[13] Scott L, Li N, Dobrev D. Role of inflammatory signaling in atrial fibrillation[J]. Int J Cardiol, 2019, 287: 195-200. DOI: 10.1016/j.ijcard.2018.10.020.
[14] Xintarakou A, Tzeis S, Psarras S, et al. Atrial fibrosis as a dominant factor for the development of atrial fibrillation: facts and gaps[J]. Europace, 2020, 22(3): 342-351. DOI: 10.1093/europace/euaa009.
[15] Lei WR, Jia LY, Wang Z, et al. CC chemokines family in fibrosis and aging: from mechanisms to therapy[J]. Ageing Res Rev, 2023, 87: 101900. DOI: 10.1016/j.arr.2023.101900.
[16] Trial J, Entman ML, Cieslik KA. Mesenchymal stem cell-derived inflammatory fibroblasts mediate interstitial fibrosis in the aging heart[J]. J Mol Cell Cardiol, 2016, 91: 28-34. DOI: 10.1016/j.yjmcc.2015.12.017.
[17] Ma J, Chen QX, Ma SY. Left atrial fibrosis in atrial fibrillation: mechanisms, clinical evaluation and management[J]. J Cell Mol Med, 2021, 25(6): 2764-2775. DOI: 10.1111/jcmm.16350.
[18] Hajam YA, Rani R, Ganie SY, et al. Oxidative stress in human pathology and aging: molecular mechanisms and perspectives[J]. Cells, 2022, 11(3): 552. DOI: 10.3390/cells11030552.
[19] Ramos-Mondragón R, Lozhkin A, Vendrov AE, et al. NADPH oxidases and oxidative stress in the pathogenesis of atrial fibrillation[J]. Antioxidants, 2023, 12(10): 1833. DOI: 10.3390/antiox12101833.
[20] Chandrasekaran A, Idelchik MDPS, Melendez JA. Redox control of senescence and age-related disease[J]. Redox Biol, 2017, 11: 91-102. DOI: 10.1016/j.redox.2016.11.005.
[21] Siddiqui MS, François M, Fenech MF, et al. Persistent γH2AX: a promising molecular marker of DNA damage and aging[J]. Mutat Res Rev Mutat Res, 2015, 766: 1-19. DOI: 10.1016/j.mrrev.2015.07.001.
[22] Mah LJ, El-Osta A, Karagiannis TC. GammaH2AX as a molecular marker of aging and disease[J]. Epigenetics, 2010, 5(2): 129-136. DOI: 10.4161/epi.5.2.11080.
[23] Talma N, Gerrits E, Wang BS, et al. Identification of distinct and age-dependent p16High microglia subtypes[J]. Aging Cell, 2021, 20(10): e13450. DOI: 10.1111/acel.13450.
[24] Adili A, Zhu XY, Cao HL, et al. Atrial fibrillation underlies cardiomyocyte senescence and contributes to deleterious atrial remodeling during disease progression[J]. Aging Dis, 2022, 13(1): 298-312. DOI: 10.14336/AD.2021.0619.
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