Xiao Qingping, Liu Lijuan, Qian Wei, Kang Ting, Ying Ru, Nie Jungang
Department of Respiratory Medicine, The First Affiliated Hospital, Jiangxi Medical College, Nanchang University, Nanchang, 330006, Jiangxi Province, People's Republic of China.
Department of Cardiology, The First Affiliated Hospital, Jiangxi Medical College, Nanchang University, No. 17, Yongwaizheng Street, Donghu District, Nanchang, 330006, Jiangxi Province, People's Republic of China.
J Cardiovasc Transl Res. 2024 Jun;17(3):540-553. doi: 10.1007/s12265-023-10478-3. Epub 2024 Jan 16.
Calcium/calmodulin-dependent protein kinase II (CaMKII) has been demonstrated to be aberrantly activated in viral myocarditis (VMC), but the role of its subtype CaMKIIδ in VMC remains unclear.VMC mice and cardiomyocytes models were induced by Coxsackievirus B3 (CVB3) treatment. Mice that underwent sham surgery and saline-treated cardiomyocytes served as controls. Body weight, survival, left ventricular ejection fraction (LVEF), and fractional shortening (LVFS) were measured, and HE staining was performed to evaluate heart function in VMC mice model and sham control. Inflammation factors in serum or cell supernatant were detected by ELISA. Expressions of CaMKIIδ, Toll/interleukin-1 receptor domain containing adaptor protein (TIRAP), insulin-like growth factor 2 mRNA binding protein 2 (IGF2BP2), nuclear factor NF-kappaB (NF-κB) signals, and inflammation factors were examined by quantitative real time polymerase chain reaction (qRT-PCR) or western blot. CCK-8, EdU, and flow cytometry were used to evaluate cell behaviors. Co-immunoprecipitation (Co-IP), RNA immunoprecipitation (RIP), and RNA pull-down were utilized to validate molecule interaction. Methylated RNA immunoprecipitation (MeRIP) was performed to measure N6-methyladenosine (m6A) level of specific molecule.CaMKIIδ was upregulated in VMC mice and CVB3-treated primary cardiomyocytes, of which knockdown improved cell viability, proliferation, and suppressed cell apoptosis in vitro, thereby alleviating myocarditis in vivo. The stability of CaMKIIδ was attributed to the presence of IGF2BP2 through m6A modification. Loss of CaMKIIδ repressed NF-κB pathway via negatively and directly regulating TIRAP to be involved in inflammatory damage.CaMKIIδ, stabilized by m6A reader IGF2BP2, modulated NF-κB pathway via interacting with TIRAP to alter cell viability, proliferation, and apoptosis, thereby affecting VMC outcome.
钙/钙调蛋白依赖性蛋白激酶II（CaMKII）已被证明在病毒性心肌炎（VMC）中异常激活，但其亚型CaMKIIδ在VMC中的作用仍不清楚。通过柯萨奇病毒B3（CVB3）处理诱导VMC小鼠和心肌细胞模型。接受假手术和生理盐水处理的心肌细胞的小鼠作为对照。测量体重、生存率、左心室射血分数（LVEF）和缩短分数（LVFS），并进行HE染色以评估VMC小鼠模型和假手术对照的心脏功能。通过ELISA检测血清或细胞上清液中的炎症因子。通过定量实时聚合酶链反应（qRT-PCR）或蛋白质免疫印迹法检测CaMKIIδ、含Toll/白细胞介素-1受体结构域的衔接蛋白（TIRAP）、胰岛素样生长因子2 mRNA结合蛋白2（IGF2BP2）、核因子NF-κB（NF-κB）信号和炎症因子的表达。使用CCK-8、EdU和流式细胞术评估细胞行为。采用免疫共沉淀（Co-IP）、RNA免疫沉淀（RIP）和RNA下拉实验验证分子间相互作用。进行甲基化RNA免疫沉淀（MeRIP）以测量特定分子的N6-甲基腺苷（m6A）水平。CaMKIIδ在VMC小鼠和CVB3处理的原代心肌细胞中上调，敲低CaMKIIδ可改善体外细胞活力、增殖并抑制细胞凋亡，从而减轻体内心肌炎。CaMKIIδ的稳定性归因于通过m6A修饰存在IGF2BP2。CaMKIIδ的缺失通过负向和直接调节TIRAP抑制NF-κB通路，从而参与炎症损伤。由m6A阅读蛋白IGF2BP2稳定的CaMKIIδ通过与TIRAP相互作用调节NF-κB通路，改变细胞活力、增殖和凋亡，从而影响VMC的结局。
