Zhu Jinlu, Liang Zijing, Yao Huochun, Wu Zongfu
MOE Joint International Research Laboratory of Animal Health and Food Safety, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing 210014, China.
Key Lab of Animal Bacteriology, Ministry of Agriculture, Nanjing 210014, China.
Antibiotics (Basel). 2024 Aug 2;13(8):725. doi: 10.3390/antibiotics13080725.
Cell-penetrating peptides (CPPs) are promising carriers to effectively transport antisense oligonucleotides (ASOs), including peptide nucleic acids (PNAs), into bacterial cells to combat multidrug-resistant bacterial infections, demonstrating significant therapeutic potential. , a Gram-positive bacterium, is a major bacterial pathogen in pigs and an emerging zoonotic pathogen. In this study, through the combination of super-resolution structured illumination microscopy (SR-SIM), flow cytometry analysis, and toxicity analysis assays, we investigated the suitability of four CPPs for delivering PNAs into cells: HIV-1 TAT efficiently penetrated cells with low toxicity against ; (RXR)XB had high penetration efficiency with inherent toxicity against ; (KFF)K showed lower penetration efficiency than HIV-1 TAT and (RXR)XB; K8 failed to penetrate cells. HIV-1 TAT-conjugated PNA specific for the essential gyrase A subunit gene (TAT-anti- PNA) effectively inhibited the growth of . TAT-anti- PNA exhibited a significant bactericidal effect on serotypes 2, 4, 5, 7, and 9 strains of , which are known to cause human infections. Our study demonstrates the potential of CPP-ASO conjugates as new antimicrobial compounds for combating infections. Furthermore, our findings demonstrate that applying SR-SIM and flow cytometry analysis provides a convenient, intuitive, and cost-effective approach to identifying suitable CPPs for delivering cargo molecules into bacterial cells.
细胞穿透肽（CPPs）是很有前景的载体，可有效地将包括肽核酸（PNAs）在内的反义寡核苷酸转运到细菌细胞中，以对抗多重耐药细菌感染，显示出巨大的治疗潜力。猪链球菌是一种革兰氏阳性菌，是猪体内主要的细菌病原体和一种新出现的人畜共患病原体。在本研究中，通过超分辨率结构光照显微镜（SR-SIM）、流式细胞术分析和毒性分析试验相结合，我们研究了四种CPPs将PNAs递送至猪链球菌细胞中的适用性：HIV-1 TAT能有效穿透猪链球菌细胞，且对其毒性较低；（RXR）XB具有高穿透效率，但对猪链球菌有内在毒性；（KFF）K的穿透效率低于HIV-1 TAT和（RXR）XB；K8未能穿透猪链球菌细胞。针对必需的gyrase A亚基基因的HIV-1 TAT偶联PNA（TAT-抗猪链球菌PNA）有效抑制了猪链球菌的生长。TAT-抗猪链球菌PNA对已知可引起人类感染的猪链球菌2、4、5、7和9型菌株表现出显著的杀菌作用。我们的研究证明了CPP-ASO偶联物作为对抗猪链球菌感染的新型抗菌化合物的潜力。此外，我们的研究结果表明，应用SR-SIM和流式细胞术分析为鉴定适合将货物分子递送至细菌细胞的CPPs提供了一种方便、直观且经济高效的方法。
