Department of Cardiology, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, China.
School of Public Management, Northwest University, Xi'an 710127, China.
Theranostics. 2023 Mar 13;13(6):1759-1773. doi: 10.7150/thno.81336. eCollection 2023.
The invasive intramyocardial injection of mesenchymal stromal cells (MSCs) allows for limited repeat injections and shows poor therapeutic efficacy against ischemic heart failure. Intravenous injection is an alternative method because this route allows for repeated, noninvasive, and easy delivery. However, the lack of targeting of MSCs hinders the ability of these cells to accumulate in the ischemic area after intravenous injections. We investigated whether and how the overexpression of colony-stimulating factor 2 receptor beta subunit (CSF2RB) may regulate the cardiac homing of MSCs and their cardioprotective effects against ischemic heart failure. Adult mice were subjected to myocardial ischemia/reperfusion (MI/R) or sham operations. We observed significantly higher CSF2 protein expression and secretion by the ischemic heart from 1 day to 2 weeks after MI/R. Mouse adipose tissue-derived MSCs (ADSCs) were infected with adenovirus harboring CSF2RB or control adenovirus. Enhanced green fluorescent protein (EGFP)-labeled ADSCs were intravenously injected into MI/R mice every three days for a total of 7 times. Compared with ADSCs infected with control adenovirus, intravenously delivered ADSCs overexpressing CSF2RB exhibited markedly increased cardiac homing. Histological analysis revealed that CSF2RB overexpression significantly enhanced the ADSC-mediated proangiogenic, antiapoptotic, and antifibrotic effects. More importantly, ADSCs overexpressing CSF2RB significantly increased the left ventricular ejection fraction and cardiac contractility/relaxation in MI/R mice. experiments demonstrated that CSF2RB overexpression increases the migratory capacity and reduces the hypoxia/reoxygenation-induced apoptosis of ADSCs. We identified STAT5 phosphorylation as the key mechanism underlying the effects of CSF2RB on promoting ADSC migration and inhibiting ADSC apoptosis. RNA sequencing followed by cause-effect analysis revealed that CSF2RB overexpression increases the expression of the ubiquitin ligase RNF4. Coimmunoprecipitation and coimmunostaining experiments showed that RNF4 binds to phosphorylated STAT5. RNF4 knockdown reduced STAT5 phosphorylation as well as the antiapoptotic and promigratory actions of ADSCs overexpressing CSF2RB. We demonstrate for the first time that CSF2RB overexpression optimizes the efficacy of intravenously delivered MSCs in the treatment of ischemic heart injury by increasing the response of the MSCs to a CSF2 gradient and CSF2RB-dependent STAT5/RNF4 activation.
间质基质细胞（MSCs）的侵袭性心肌内注射允许有限的重复注射，并显示出对缺血性心力衰竭的治疗效果不佳。静脉内注射是一种替代方法，因为这种途径允许重复、非侵入性和易于递送。然而，MSCs 的靶向缺乏阻碍了这些细胞在静脉内注射后在缺血区域的积累能力。我们研究了过表达集落刺激因子 2 受体β亚基（CSF2RB）是否以及如何调节 MSCs 的心脏归巢及其对缺血性心力衰竭的心脏保护作用。成年小鼠接受心肌缺血/再灌注（MI/R）或假手术。我们观察到，从 MI/R 后 1 天到 2 周，缺血心脏的 CSF2 蛋白表达和分泌显著增加。用携带 CSF2RB 或对照腺病毒的腺病毒感染小鼠脂肪组织源性 MSCs（ADSCs）。增强型绿色荧光蛋白（EGFP）标记的 ADSC 每 3 天静脉注射一次，共 7 次。与感染对照腺病毒的 ADSC 相比，静脉内递送过表达 CSF2RB 的 ADSC 表现出明显增加的心脏归巢。组织学分析显示，CSF2RB 过表达显著增强了 ADSC 介导的促血管生成、抗凋亡和抗纤维化作用。更重要的是，过表达 CSF2RB 的 ADSC 显著增加了 MI/R 小鼠的左心室射血分数和心脏收缩/松弛。实验表明，CSF2RB 过表达增加了 ADSC 的迁移能力，并减少了缺氧/复氧诱导的 ADSC 凋亡。我们确定 STAT5 磷酸化是 CSF2RB 促进 ADSC 迁移和抑制 ADSC 凋亡的关键机制。RNA 测序和因果分析表明，CSF2RB 过表达增加了泛素连接酶 RNF4 的表达。共免疫沉淀和共免疫染色实验表明，RNF4 与磷酸化的 STAT5 结合。RNF4 敲低减少了 STAT5 磷酸化以及过表达 CSF2RB 的 ADSC 的抗凋亡和促迁移作用。我们首次证明，CSF2RB 过表达通过增加 MSCs 对 CSF2 梯度的反应和 CSF2RB 依赖性 STAT5/RNF4 激活，优化了静脉内递送 MSCs 治疗缺血性心脏损伤的疗效。
