Ainiwan Ailifeire, Wei Yuanyuan, Dou Jing, Tang Lingpeng, Mu Yuming, Guan Lina
Department of Echocardiography, The First Affiliated Hospital of Xinjiang Medical University, Urumqi, China.
Front Med (Lausanne). 2023 Sep 22;10:1136304. doi: 10.3389/fmed.2023.1136304. eCollection 2023.
Stem cell transplantation is one of the treatment methods for acute myocardial infarction (AMI). MicroRNA-1 contributes to the study of the essential mechanisms of stem cell transplantation for treating AMI by targeted regulating the myocardial microenvironment after stem cell transplantation at the post-transcriptional level. Thus, microRNA-1 participates in regulating the myocardial microenvironment after stem cell transplantation, a promising strategy for the Stem cell transplantation treatment of AMI. However, the naked microRNA-1 synthesized is extremely unstable and non-targeting, which can be rapidly degraded by circulating RNase. Herein, to safely and effectively targeted transport the naked microRNA-1 synthesized into myocardial tissue, we will construct pseudo-endogenous microRNA-targeted myocardial ultrasound nanobubble pAd-AAV-9/miRNA-1 NB and evaluate its characteristics, targeting, and function.
The pAd-AAV-9/miRNA-1 gene complex was linked to nanobubble NBs by the "avidin-biotin bridging" method to prepare cardiomyocyte-targeted nanobubble pAd-AAV-9/miRNA-1 NB. The shape, particle size, dispersion, and stability of nanobubbles and the connection of pAd-AAV-9/miRNA-1 gene complex to nanobubble NB were observed. The virus loading efficiency was determined, and the myocardium-targeting imaging ability was evaluated using contrast-enhanced ultrasound imaging . The miRNA-1 expression level in myocardial tissue and other vital organs of SD rats was considered by Q-PCR. Also, the cytotoxic effects were assessed.
The particle size of NBs was 504.02 ± 36.94 nm, and that of pAd-AAV-9/miRNA-1 NB was 568.00 ± 37.39 nm. The particle size and concentration of pAd-AAV-9/miRNA-1 NBs did not change significantly within 1 h at room temperature ( > 0.05). pAd-AAV-9/miRNA-1 NB had the highest viral load rate of 86.3 ± 2.2% ( < 0.05), and the optimum viral load was 5 μL ( < 0.05). pAd-AAV-9/miRNA-1 NB had good contrast-enhanced ultrasound imaging . Quantitative analysis of miRNA-1 expression levels in vital organs of SD rats by Q-PCR showed that pAd-AAV-9/miRNA-1 NB targeted the myocardial tissue. Q-PCR indicated that the expression level of miRNA-1 in the myocardium of the pAd-AAV-9/miRNA-1 NB + UTMD group was significantly higher than that of the pAd-AAV-9/miRNA-1 NB group ( < 0.05). pAd-AAV-9/miRNA-1 NB had no cytotoxic effect on cardiomyocytes ( > 0.05).
The pAd-AAV-9/miRNA-1 NB constructed in this study could carry naked miRNA-1 synthesized for targeted transport into myocardial tissue successfully and had sound contrast-enhanced imaging effects .
干细胞移植是急性心肌梗死（AMI）的治疗方法之一。微小RNA-1通过在转录后水平靶向调控干细胞移植后心肌微环境，有助于研究干细胞移植治疗AMI的关键机制。因此，微小RNA-1参与调控干细胞移植后的心肌微环境，这是干细胞移植治疗AMI的一种有前景的策略。然而，合成的裸微小RNA-1极其不稳定且无靶向性，会被循环中的核糖核酸酶迅速降解。在此，为了将合成的裸微小RNA-1安全有效地靶向转运至心肌组织，我们将构建伪内源性微小RNA靶向心肌超声纳米泡pAd-AAV-9/miRNA-1 NB，并评估其特性、靶向性和功能。
采用“抗生物素蛋白-生物素桥接”法将pAd-AAV-9/miRNA-1基因复合物与纳米泡NB相连，制备心肌靶向纳米泡pAd-AAV-9/miRNA-1 NB。观察纳米泡的形状、粒径、分散性和稳定性以及pAd-AAV-9/miRNA-1基因复合物与纳米泡NB的连接情况。测定病毒装载效率，并使用超声造影成像评估心肌靶向成像能力。通过Q-PCR检测SD大鼠心肌组织及其他重要器官中miRNA-1的表达水平。同时，评估细胞毒性作用。
NB的粒径为504.02±36.94nm，pAd-AAV-9/miRNA-1 NB的粒径为568.00±37.39nm。pAd-AAV-9/miRNA-1 NBs的粒径和浓度在室温下1小时内无明显变化（>0.05）。pAd-AAV-9/miRNA-1 NB的病毒装载率最高，为86.3±2.2%（<0.05），最佳病毒装载量为5μL（<0.05）。pAd-AAV-9/miRNA-1 NB具有良好的超声造影成像效果。通过Q-PCR对SD大鼠重要器官中miRNA-1表达水平进行定量分析，结果表明pAd-AAV-9/miRNA-1 NB靶向心肌组织。Q-PCR显示，pAd-AAV-9/miRNA-1 NB+UTMD组心肌中miRNA-1的表达水平显著高于pAd-AAV-9/miRNA-1 NB组（<0.05）。pAd-AAV-9/miRNA-1 NB对心肌细胞无细胞毒性作用（>0.05）。
本研究构建的pAd-AAV-9/miRNA-1 NB能够成功携带合成的裸miRNA-1靶向转运至心肌组织，并具有良好的超声造影成像效果。
